Lincomycin derivatives and process for preparing same



United States Patent 3,366,624 LINCOMYCHN DERHVATIVES AND PROCESS FORPREPARING SAME Alexander D. Argoudeiis and Brian Bannister, Kalamazoo,Herman Hoeksema, Cooper Township,-Kalamazoo County, and Fred Kagan andBarney J. Magerlein, Kalamazoo, Mich, assignors to The Upjohn Company,Kalamazoo, Mich, a corporation of Delaware No Drawing. Filed Aug. 1964,Ser. No. 387,776 14 Claims. (Cl. 260-210) This invention relates to amethod of producing lincomycin and analogs and isomers thereof and tonew compounds produced thereby.

Lincomycin is an antibiotic obtained as an elaboration product of alincomycin-producing actinomycete. Methods for the production, recovery,and purification of lincomycin are described in US. Patent 3,086,912.Partly as a result of this invention the structure of lincomycin hasbeen elucidated as methyl 6-(trans-4-propyl-l-methyl- L-prolylamino)6,8-dideoxy 1 thio-D-eryth.ro a D- galacto-octopyranoside which has thefollowing structural formula:

( ino It has now been found that lincomycin and isomers and analogsthereof can now be synthesized by acylating with a 4-substitutedL-proline of the formula:

il-OH OH tj Y I CIR; III wherein R and R are hydrogen or alkyl of notmore than 12 carbon atoms and wherein Y is hydrogen, 3 alkyl of not morethan 12 carbon atoms, and

SCH CH OR wherein R is hydrogen, or alkyl of not more than 12 carbonatoms, to form compounds of the formula:

3,356,624 Patented Jan. 36, 1968 wherein Z, Y, R, R and R are as givenabove; removing the radical Z by hydrogenolysis; and, if desired,alkylating the resulting compound (Formula V Wherein R is hydrogen) toproduce a compound of the formula:

0R V wherein R is alkyl of not more than 20' carbon atoms, cycloalkyl offrom three to not more than 8 carbon atoms, or aralkyl of not more than12 carbon atoms and Y, R, R and R are as given above.

Examples of alkyl of not more than 20 carbon atoms are methyl, ethyl,propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, undecyl,dodecyl, tridecyl, tetradecyl, pentadecyl, hexadecyl, heptadecyl,octadecyl, nonadecyl, and eicosyl and the isomeric forms thereof.Examples of cycloalkyl are cyclopropyl, cyclobutyl, cyclopentyl,cyclohexyl, cycloheptyl, cyclooctyl, Z-methylcyclopentyl,2,3-dimethylcyclobutyl, 4-methylcyclobutyl, and 3-cyclopentylpropyl.Examples of aralkyl are benzyl, phenethyl, a-phenylpropyl, anda-naphthylmethyl. Examples of hydrocarboxycarbonyl groups aretertiary-butoxycarbonyl; benzyloxycarbonyl groups of the formula:

wherein X is hydrogen, nitro, methoxy, chl-oro or bromo, for example,carbobenzoxy, p-nitrocarbobenzoxy, pbromo, and p-chlorocarbobenzoxy; andphenyloxycarbonyl groups of the formula:

wherein X is hydrogen, allyl or alkyl of not more than carbonyl,p-ethylphenyloxycarbonyl, and allyloxycarbonyl and the like.

The starting compounds of Formula II are prepared by thefollowingsequence:

0 01-1 C-OII 11* 0 It I 41 VI VII r! i N I COH A H H an 0 The resultingproduct is a mixture of the cis and trans isomers.

Ila III) N N 1 1 \I RH Lw (fi-ou t p-0n Cis The above sequence iseffected by treating a 1 hydrocarbyloxycarbonyl 4 keto L proline (VI),or otherwise protected 4 keto L proline, with a Wittig reagent, usuallyan alkylidenetriphenylphosphorane [see e.g., Wittig et 211., Ben, 87,1,348 (1954); Trippett, Quarterly Reviews, XVII, No. 4, p. 406 (1963)],and hydrogenating the obtained 4 alkylidene 1 hydrocarbyloxycarbonyl- Lproline (VII) in the presence of a platinum catalyst to obtain thecorresponding 4 alkyl l hydrocarbyloxycarbonylproline (II).

In carrying out the process the 1 hydrocarbyloxycarbonyl 4 keto Lproline (V1), is added to a freshly prepared Wittig reagent. The Wittigreagents herein used can be generally represented by the followingformula:

wherein R is alkylidene of not more than 12 carbon atoms,cycloalkylidene of from 3 to not more than 8 carbon atoms, andaralkylidene of not more than 12 carbon atoms. Suitable alkylidene,cycloalkylidene, and aralkylidene groups include methylene, ethylidene,propylidene, butylidene, pentylidene, hexylidene, heptylidene,octylidene, nonylidene, decylidene, undecylidene, and dodecylidene andthe isomeric forms thereof, cyclopropylidene, cyclobutylidene,cyclopentylidene, cyclohexylidene, cycloheptylidene, cyclooctylidene,2-cyclopropylethylidene, 3- cyclohexylpropylidene, benzylidene,Z-phenylethylidene, 3-phenylpropylidene, and a-naphthylmethylene. TheseWittig reagents are prepared by reacting an alkylcyclo alkyl, oraralkyltriphenylphosphonium halide with a base such as sodamide, orsodium or potassium hydride, or the sodium or potassium metalate ofdimethylsulfoxide and the like. For example, the elimination of hydrogenhalide from alkyltriphenylphosphoniurn halide, producesalkylidenetriphenylphosphorane. [The preparation of phosphoranes isdiscussed in detail by Trippett, Quart. Rev. XVII, No. 4, p. 406(1963).] The reaction is generally carried out in an organic solvent,such as benzene, toluene, ether, dimethylsulfoxide, tetrahydrofuran, orthe like, at temperatures between 10 C. and the reflux temperature ofthe reaction mixture. The thus-obtained product, a 4-alkylidene-,4-cycloalkylidene-, or 4-aralkylidene-l-hydrocarbyloxycarbonyl L proline(VII), is recovered from the the reaction mixture in a conventionalmanner, generally by extraction from aqueous solutions of the reactionmixture. The crude product can be purified by conventional means, suchas recrystallization, chromatography, or formation and recrystallizationof easily formed derivatives such as amine salts of the amino acid,e.g., the dicyclohexylamine salt, and the like, and liberating the aminoacids from such compounds.

The thus-obtained 4 alkylidene-, 4 cycloalkylidene, or 4 aralkylidene 1hydrocarbyloxycarbonyl L proline (VII) is then hydrogenated in thepresence of platinum, which is generally deposited on a carrier forexample, carbon or an anion exchange resin like Dowex-l, a cross-linkedpolystyrene trimethylbenzylammonium resin. A mixture of the cisandtrans-epimers of the l-hydrocarbyloxycarbonyl-4-substituted-L-proline(Ila and 11b) is obtained.

Alternatively, the process can be carried out by acylating Compound IIIwith Compound VII to form a compound of the formula:

and then hydrogenating the alkylidene group with a platinum or palladiumcatalyst. With a platinum catalyst as given above the Z radical ispreserved but with a palladium catalyst, e.g., palladium on carbon,hydrogenolysis of the Z group concurrently with saturation of thealkylidene group is obtained.

The starting compounds of Formula III can be obtained in a variety ofways. Methyl 6 amino 6,8 dideoxy l thio D erythro a D galactooctopyranoside, also designated as methyl a-thiolincosaminide or MTL, isobtained by subjecting lincomycin to hydrazinolysis and ethyl 6 amino6,8 dideoxy 1 thio D erythro 0c D galacto octypyranoside, also known asethyl 0:thi0- lincosaminide or ETL, is obtained by hydrazinolysis oflincomycin C, an antibiotic obtained when the lincomycin fermentation ofU.S. Patent 3,086,912 is carried out in the presence of ethionine.

The hydrazinolysis advantageously is effected by heating the antibioticunder reflux with an excess of hydrazine, say for 20 hours or more. Thedesired 6-amino-6,8-dideoxy 1 thio D erythro a D galacto octopyranosidecan be isolated by distilling off the excess hydrazine and crystallizingfrom a polar solvent such as ethanol.

Other 6 amino 6,8 dideoxy 1 thio D erythro a D galacto octopyranosidescan be prepared by the following representative sequences:

Ac is alkanoyl or aralkanoyl of not more than 12 carbon atoms. Examplesare formyl, acetyl, propionyl, butyryl, valeryl, hexanoyl, heptanoyl,octanoyl, nonanoyl, decanoyl, undecanoyl, dodecanoyl, and the isomericforms thereof, and benzoyl, Z-phenylacetyl, 3-phenylpropionyl, 4-phenylbutyryl, S-p'henylvaleryl and the isomeric forms 7 thereof.

The process is effected as follows: Methyl a-thiolincosaminide isN-acylated, the methyl N-acyl-oc-thiolincosaminide (IX) then O-acylatedwith thiobenzoyl chloride, and the resulting thiobenzoyl ester (X) isbrominated to form Compound XI which on treatment with a base isconverted to Compound XII. Treatment of Compound XII with mild alkali(aq. carbonate-bicarbonate) gives Compound XIII which on alkylation withan alkyl halide yields Compound XIV. Hydrolysis of the benzoyl group andhydrazinolysis of the N-acyl group gives Compound IIIb. In a like mannercompounds in which Y is Z-hydroxyethyl or 2-alkoxyethyl can be preparedby reacting Compound XIII with ethylene chlorhydrin or a 2-alkoxyethylhalide.

The ,B-epimers of the above can be prepared by the followingrepresentative sequence:

Sequence B CIIg I-IO

AcNH

methyl N -acy1-athiolincosaminide Ac and Ac can be alkanoyl oraralkano-yl of not more than 12 carbon atoms as given above.

The process is effected as follows: treating methyl N-acyl-a-thiolincosaminide (IX) with mercuric chloride in a warm aqueoussolution to obtain a mixture of 6-acy1-amino-6,8-dideoxy-D-erythro-a-D-galacto octopyranose orN-acyl-a-lincosamine (XVot) and N-acyl-fi-lincosamine (XV/3); acylatingthis mixture with an acylating reagent selected from acyl halides andacid anhydrides to obtain a mixture ofN-acyl-l,2,3,4,7-penta-O-acyl-aand ,B-lincosamine (XVIa and XVI/3);treating Compound XVIoc or Compound XVIB or a mixture of Compounds XVICZand XVIB with hydrogen bromide in acetic acid to obtain 6acylamino-2,3,4,7-tetra-O-acyl-la-bromo-1,6,8-trideoXy-D-erythro-D-galacto-octopyranoseor N-acy1-2,3,4, 7 tetra O acyl-la-bromo-l-deoxylincosamine (XVII);treating Compound XVII successively with thiourea, a mixture ofpotassium carbonate and sodium bisulfite in water, and an alkyl iodideto obtain alkyl N-acyl-2,3,4,7- tetra-O-acyl-/3-thiolincosaminide(XVIII); treating Compound XVIII with dry ammonia gas in methanol toobtain alkyl N-acyl-B-thiolincosaminide (XIX). Hydrazinolysis ofCompound XIX or of Compound XVIII gives the alkyl ti-thiolincosaminide(IIIc). In a like manner compounds in which Y (Formula III) is2-hydroxyethylthio or 2-alkoxyethylthio can be prepared by reactingCompound XVII with ethylenechlorhydrin or a 2-alkoxyethyl halide.

Compounds of Formula III where Y is hydrogen can be obtained by thefollowing representative sequence:

The process is carried out as follows: methyl N-acyLathiolincosaminide(IX) (or methyl N-acyl-fi-triolincosaminide) is treated with acetone inthe presence of sulfuric acid to form the methylN-acyl-3,4-O-isop-ropylidene-athiolincosaminide (XX) which can bedesulfurized with Raney nickel in ethanol to 6-acylamino-3,4O-isopropylidene-1,6,8-trideoxy-D-erythro-D-galacto-octopyranose, or Nacyl-3,4-O-isopropylidene-l-deoxylincosamine (XXI). The desulfurizationadvantageously is achieved by heating under reflux in ethanol withloosely packed Raney nickel for a period of 2 to 24- hours. Thereafter,the catalyst is removed by filtration and the filtrate is evaporated togive a residue which canbe purified by conventional means such asrecrystallization from organic solvents. The isopropylidene group can beremoved by mild acid hydrolysis, e.g., aqueous acetic acid or a dilutemineral acid such as hydrochloric acid or sulfuric acid, to giveNacyl-1-deoxylincosamii1e (XXII) and the N-acyl by hydrazinolysis togive l-deoxylincosarnine (XIII). Al-

ternatively the hydrazinolysis can be effected first to give 3,4 Oisopropylidene-l-deoxylincosamine (XXIV). Advantageously, however, theisopropylidene group is removed first.

Starting D-erythro-D-galacto-compounds of Formula III wherein R and/or Ris alkyl of not more than 12 carbon atoms can be obtained by thefollowing representative sequence:

Sequence D can be alkanoyl or aralkanoyl of not more than 12 carbonatoms as given above. R and R can be different alkyls if CompoundsXXVIIla and/ or XXVIITb are alkylated with a diiferent alkylating agentfrom that used to alkylate compound XXVII. R and R in Compound XXVIIIccan thus be different alkyls.

The process is effected as follows: treating a 6-acylamino 6,8dideoxy-D-erythro-D-galacto-octopyranose compound of Formula XXV withdry acetone in the presence of an acid catalyst, for example, sulfuricacid, to give a mixture of Compound XXVII and the oxazoline derivativethereof XXVI; hydrolyzing Compound XXVI in hot water to convert it toCompound XXVII; methylating Compound XXVII with an alkyl halide, forexample alkyl chlorides, alkyl bromides, or alkyl iodides of not morethan 12 carbon atoms, in the presence of a base and separating the threeproducts thus obtained (Compound XXVIIIa, Compound XXVIIIb, and CompoundXXVIIIc); and hydrolyzing the Compound XXVIII under mild hydrolyzingconditions, for example, with 80% aqueous acetic acid or a dilutemineral acid, such as hydrochloric acid or sulfuric acid, to eliminatethe isopropylidene moiety and treating with hydrazine to remove theN-acyl group to give the compound XXIX. When Y is Z-hydroxyethylthio inthe above sequence or when 2- hydroxyethyl 6amino-6,8-dideoxy-7-O-methyl-l-thio-D-erythro-a-D-galacto-octopyranoside, also known as 2-hydroxyethyla-thiocelestosaminide, obtained by the hydrazinolysis of celesticetin,an antibiotic produced according to US. Patent 2,928,844, is used, the2-hydroxy group will also be alkylated to give a 2-alkoxyethylthio groupin which the alkoxy group will contain not more than 12 carbon atomsaccording to the alkyl halide used to alkylate. When desired the2-hydroxythio group can be preserved by protecting it with a tritylgroup. Thus Compounds XXV or XXV-II where Y is 2-hydroxyethylthio can bereacted with trityl chloride (triphenylmethylchloride) chlorodiphenyl(p-methoxyphenyl)methane, or chloro-bis-(p-methoxyphenyl)phenylmethane,to convert Y to a 2-trityloxyethylthio and the trityl group removed bymild hydrolysis, for example with aqueous acetic acid, after thealkylation. The 2-hydroxy group can be alkylated with a different alkylgroup.

In carrying out this process 6-acylamino-6,8-dideoxy-D-erythro-D-glacto-octopyranose compound of Formula XXV is suspended indry acetone. An acid catalyst, such as sulfuric acid, p-toluenesulfonicacid, o-toluenesulfonic acid, p-ethylbenzenesulfonic acid, or the like,is added to the suspension, with sulfuric acid preferred. The acetone isgenerally used in large excess, such as a volume of 20 to 200 times theamount of Compound XXV. The acid catalyst is used in a quantity between0.25 and 5% by weight with respect to the acetone. The reaction can becarried out between 10 C. and the reflux temperature of the solution,but is generally carried out at room temperature. The reaction time isbetween 15 minutes and 6 hours, depending on the temperature, afterwhich the reaction mixture is neutralized to terminate the reaction. Theinorganic salts precipitated by the neutraliztaion are removed byfiltration, and the filtrate is evaporated to give a mixture containingcrystalline solids. This mixture is separated with water into awater-soluble and water-insoluble fraction; the water-soluble materialis Compound XXVII; the insoluble fraction is the oxazoline derivativethereof (XXVI).

The water-insoluble oxazoline derivative (XXVI) can be converted toCompound XXVII by heating with water. After this hydrolysis is complete,the product is obtained by evaporating the solution untilcrystallization occurs.

The alkylation of Compound XXVII is performed with an alkyl halide,preferably an alkyl chloride, bromide or iodide, in the presence of astrong base. As alkylating agents, methyl iodide, methyl bromide, ethyliodide, ethyl bromide, and propyl, butyl, isobutyl, pentyl, hexyl,heptyl, oc-tyl, nonyl, decyl, undecyl, dodecyl chlorides, bromides andiodides and any of the branched-chain alkyl chlorides, bromides oriodides having up to 12 carbon atoms can be used. As a base, alkoxides,such as sodium methoxide, potassium methoxide, potassium isopropoxide,potassium tert-butoxide, sodium tert-butoxide, and soda-mide and thelike can be used. The alkylation can be performed at a temperaturebetween 0 and 50 C. using inert solvents such as benzene, toluene,dioxane, tetrahydrofuran, and the like. After the reaction isterminatedusually between /2 hour and 3 hours-the reaction mixture isfiltered'to remove solids, such as sodium or potassium chloride, bromideor iodide, and the filtrate is evaporated in vacuo to a syrupy residue.The three products therein, namely Compound XXVIIIa, Compound XXVIIIb,and Vompound XXVIIIc, are separated by conventional methods, forexample, by chromatography or by counter-current distribution. Theremoval of the N-acyl and isopropylidene groups is effected as describedabove for sequence C. It will be understood that 2-0-, and 7-0, and2,7-di-O-alkylates according to Formulas XXIXa, XXIXb, and .XXIXc can beused in Sequences A, B, and C to give the 2-0, 7-0, and2,7-di-O-alkylates of compounds produced in these sequences.

The 7-O-epimers of the Compounds III, i.e., the 6-amino-6,8-dideoxy-L-threo-D-galacto octopyranose compounds, can beobtained by using as the starting compound epi MTL (methyl6-amino-6,8-dideoxy-1-thio-L threo-e-D-galacto-octopyranoside) havingthe formula:

The epi-MTL can be prepared by the following representative sequence:

Ac can be alkanoyl or aralkanoyl of not more than 12 carbon atoms asgiven above or trans-4-propyl-l-methyl- L-prolyl in which case IX islincomycin (I) and XXXII is epilincomycin [6-(trans 4propyl-l-methyl-L-prolylamino)-6,8-dideoxy1-thio-L-threo-a-D-galacto-octopyranoside].

Alternatively the above sequence can be applied to any of the6-amino-6,8-dideoxy-D-erythro-D-galacto octopyranose compounds ofFormula III (or N-acylates thereof) disclosed above.

In carrying out the process the star-ting compound is an N-acylderivative of a 6-amino-6,8-dideoXy-D-erythro- -galacto-octopyranosecompound of Formula III which advantageously is lincomycin. Thus in anillustrative procedure lincomycln (I) is reacted with acetone to formthe 3,4-O-isopropylidene derivative XXa by the procedure described abovefor Sequences C and D. As the starting compound is lincomycin formationof the oXazoline XXVI is minimized. The 3,4-O-isopropylidene-lincomycin,XXa, is then oxidized with chromic acid to form the 7-dehydroderivative, XXX, which in turn is reduced with borohydride to a mixtureof 3,4-O-isopropylidene-lincomycin and3,4-0-isopropylidene-epilincomycin (XXXI). The two epimers can beseparated by fractional liquid-liquid extraction, for example,counter-current distribution or partition chromatography, or byadsorption or gradient chromatography, or the mixture can be convertedto a mixture of lincomycin (I) and epilincomycin (XXXII) and the twoepimers then separated by like procedures. Removal of the3,4-O-isopropylidene group can be effected by mild hydrolysis asdescribed above for Sequence C, to give epilincomycin (XXXII) which inturn can be deacylated with. hydrazine as described above to form thedesired methyl epi-a-thiolincosaminide (methyl 6-amino- 6,8-dide-oxy 1thio-L-threo-w-D-galacto-octopyranoside) or the hydrogenoiysis can beetfected first and the re moval of the isopropylidene group last asdescribed for Sequence C.

Removal of the protective group Z in compounds of Formula 1V is effectedby hydrogenolysis using a palladium catalyst. The palladium is usuallydeposited on a carrier, for example, carbon. Any of the conventionalmethods of hydrogenolysis can be used.

The proline nitrogen in compounds of Formula V where R, is hydrogen canbe alkylated with an alkyl halide, for example, an alkyl iodide. Milderconditions than those given above for alkylating the 2-0 and7-O-positions are used. Thus the N-alkylation can be effected without astrong base and at room temperature or at most with mild heating, say,to not more than C. or so. Advantageously the alkylation is effected byreacting the compound with a ketaldone (an aldehyde or a ketone) andhydrogenating the resulting adduct. The hydrogenation can be efiectivewith palladium or platinum as the catalyst or any hydrogenating catalysteffective to saturate an olefinic double bond. Suitable ketaldones havethe formula: R R CO Where R R C= is alkylidene of not more than 20carbon atoms, cycloalkylidene of from 3 to not more than 8 carbon atoms,or aralkylidene of not more than 12 carbon atoms. Examples areformaldehyde,

acetaldehyde,

propionaldehyde, butyraldehyde,

acetone,

isobutyl methyl ketone, benzaldehyde, phenylacetaldehyde,hydrocinnamaldehyde, acetophenone,

propionphenone, butyrophenone, 3-methyl-4-phenyl-2-butanone,2-methyl-5-phenyl-3-pentanone, 3-cyclopentanepropionaldehyde,cyclohexaneacetaldehyde, cycloheptanecarboxaldehyde,2,2-dimethylcyclopropylacetaldehyde, 2,2-dimethylcyclopropyl methylketone, cyclopentyl methyl ketone, cyclobutyl methyl ketone,cyclobutanone,

.cyclohexanone,

4-methylcyclohexanone, and the like.

The cis and trans epimers can be separated by partition or gradientchromatography. The separation can be effected most advantageously atthe stage represented by Formulas IV, or V. The respective epimers canalso be formed by starting with the trans or cis forms of the4-substituted- L-prolines (Formulas Ila and 11b.)

Various acid-addition salts of the free base form of the compounds ofFormula V wherein R is hydrogen, alkyl, cycloalkyl, or aralkyl as givenabove, can be made by neutralizing the free base with the appropriateacid to below about pH 7.0, and advantageously to about pH 2 to pH 6.Suitable acids for this purpose include hydrochloric, sulfuric,phosphoric, thiocyanic, fluosilicic, heXafiuoroarsenic,hexafluorophosphoric, acetic, succinic, citric, lactic, maleic, fumaric,parnoic, cholic, palmitic, mucic, camphoric, glutaric, glycolic,phthalic, tartaric, lauric, stearic, salicylic, 3-phenylsalicylic,S-phenylsalicylic, 3- methylglutaric, orthosulfobenzoic,cyclohexanesulfamic, cyclopentanepropionic, 1,2-cyclohexanedicarboxylic,4-cyclohexenecarboxylic, octadecenylsuccinic, octenylsuccinic,methanesulfonic, benzenesulfonic, helianthic, Reineckes,dimethyldithiocarbamic, cyclohcxylsulfamic, hexadecylsul- 1 l famic,octadecylsulfamic, sorbic, monochloroacetic, undecylenic,4-hydroxyazobenzene-4-sulfonic, octyldecylsulfuric, picric, benzoic,cinnamic, and like acids.

The acid-addition salts can be used for the same purposes as the freebase or they can be employed to upgrade the same. For example, the freebase can be converted to an insoluble salt, such as the picrate, whichcan be subjected to purification procedures, for example, solvent extractions and washings, chromatography, fractional liquidliquidextractions, and cyrstallization and then used to regenerate the freebase form by treatment with alkali or to make a different salt bymetathesis. Or the free base can be converted to a water-soluble salt,such as the hydrochloride or sulfate and the aqueous solution of thesalt extracted with various water-immiscible solvents beforeregenerating the free base form by treatment of the thusextracted acidsolution or converted to another salt by metathesis.

The compounds of Formula V wherein R is hydrogen, alkyl, cycloalkyl, oraralkyl as given above, can be used as a buffer or as an antacid. Thecompounds of Formulas IV and V react with isocyanates to form urethanesand can be used to modify polyurethane resins. The long chain compounds,i.e., where R is alkyl of from 8 carbon atoms up, have surface activeproperties and can be used as wetting and emulsifying agents. Thethiocyanic acid addition salt when condensed with formaldehyde formsresinous materials useful as pickling inhibitors according to U.S.Patents 2,425,320 and 2,606,155. The free bases also make good vehiclesfor toxic acids. For example, the fiuosilicic acid addition salts areuseful as mothproofing agents according to U.S. Patents 1,915,334 and2,075,359 and the hexafluoroarsenic acid and hexafluorophosphoric acidaddition salts are useful as parasiticides according to US. Patents3,122,536 and 3,122,552.

The close analogues of lincomycin, i.e., where RH- is transalkyl of notmore than 4 carbon atoms; R is methyl or ethyl; Y is a-alkylthio of notmore than 4 carbon atoms have antibacterial properties comparable tolincomycin and can be used for the same purposes as lincomycin. Theother analogues and isomers have similar antibacterial properties but toa lesser degree and can be used for the same purposes as lincomycinwhere larger amounts are not objectionable.

The following examples are illustrative of the process and products ofthe present invention but are not to be construed as limiting. The partsand percentages are by weight and the solvent ratios are by volumeunless otherwise specified.

Example 1.--Methyl 6-(cisand trans-4-blltyl-1-alkyl-L- prlylamin0)-6,8dideoxy-J-thio-D-erythr0-a-D-gn[actooctopyranoside (methyl N (cisandtrans-4-buZyl-1- alkyl-L-prolyl)-e-thi0linc0saminide) XXXIV Ri=alkyl A.4-BUTYLIDENE-LCARBOBENZOXY-L-PROLINE AND THE CYCLOHEXYLAMINE SALTTHEREOF Sodium hydride (19 g.) as a 53% suspension in mineral oil waswarmed with 350 ml. of dimethylsulfoxide at a temperature of 70-75 C.until the reaction was complete (about 30 minutes). After cooling to 32C., 16.2 g. of

butyltriphenylphosphonium bromide was added, and the resulting reactionmixture was stirred for 1 hour to insure complete reaction. A solutionof 26 g. of 4-keto-l-carbobenzoxy-L-proline in ml. of dimethylsulfoxidewas added, and the resulting mixture was heated at 70 C. for 3 hours.The reaction mixture was cooled to 25 C. and 1 liter of 2.5% aqueouspotassium bicarbonate added. This mixture was washed twice with 700I111. portions of ether and the ether was discarded after backextracting with ml. of 2.5% aqueous potassium bicarbonate. Thebicarbonate solutions were combined and acidified with 4 N hydrochloricacid. The acidified mixture was extracted with four 500-ml. portions ofether. The combined ether extracts were Washed successively with 250 ml.of water, three 250-ml. portions of saturated aqueous sodium bisulfite,and 250 ml. of water, and dried over anhydrous sodium sulfate.Evaporation of the solvent under vacuum gave 24 g. of an oily residuewhich was 4-butylidene-1-carbobenzoxy-L-proline.

This residue was dissolved in 31 ml. of acetonitrile and treated with 18ml. of dicyclohexylamine and refrigerated. The crystals were collected,washed with acetonitrile and dried in vacuo giving 21 g. (46.8%) of thecrystalline dicyclohexylamine salt melting at 136140 C. After tworecrystallizations from acetonitrile, an analytical sample was obtainedwhich melted at 142144 C. and had a rotation of [M -4 (c.=0.99, CHClAnalysis.Calcd. for C l-i N O C, 71.86; H, 9.15; N, 5.78. Found: C,71.69; H, 9.30; N, 5.74.

Ten grams of the dicyclohexylamine salt of4-butylidene-l-carbobenzoxy-L-proline was shaken with ether and excess5% aqueous potassium hydroxide until no solid remained. The layers wereseparated and each one was backwashed. The aqueous alkaline layer wascombined with the backwash from the ether layer and acidified with 4 Nhydrochloric acid. The mixture was repeatedly extracted with ether andthe ether extracts were combined, dried over sodium sulfate, andevaporated in vacuo to give 6.3 g. (93%) of 4-butylidene-l-carbobcnzoxyL- proline as an oil.

13. LBUTYLl-CARBOBENZOXY-L-PROLINE The oil from Part A was hydrogenatedin 200 ml. of methanol over 2.1 g. of 10% platinum on Dowex-l catalystunder 40 lbs. hydrogen pressure. The catalyst was removed by filtrationand the filtrate evaporated to yield 6.3 g. of4-butyl-1-carbobenzoxy-L-proline as an oil. The product contained about2 parts cis-4-butyl-1-carbobenzoxyL-proline to each part oftrans-4-butyl-1-carbobenzoxy-L-proline.

C. METHYL (i AMINO-6,SDIDEOXY-1-THIO-D-ERYTHRO- a DGALACTO-OCTOPYRANOSIDE (METHYL a-THIO- LINCOSAMINIDE) A solution of 40g. of lincomycin (U.S. Patent 3,086,- 912) in 20 ml. of hydrazinehydrate (98l00%) was refluxed for 21 hours; excess hydrazine hydrate wasthen removed in vacuo under nitrogen at steam bath temperature, leavinga residue. The residue, a pasty mass of crystals, was cooled,acetonitrile was added, and the mixture was stirred until the crystalswere suspended. The crytsals were collected on a filter, washed withacetonitrile and with ether. The yield of white, crystalline methyla-thiolincosaminide after drying in va-cuo at room temperature was 21 g.(84%). Recrystallization was accomplished by dissolving methylu-thiolincosaminide in hot dimethylformarnide and adding an equal volumeof ethylene glycol dimethyl ether.

Methyl a-thiolincosaminide has a melting point of 225- 228 C., anoptical rotation of [a] +276 (c.=.768, water) and a pKa' of 7.45.

Analysis.-Calcd. for C H NO S: C, 42.7; H, 7.56; N, 5.53; S, 12.66.Found: C, 42.6; H, 7.49; N, 5.75; S, 12.38.

t t N N lhTL I/ coon giy c on'rr To a solution of 6.3 g. of4-butyl-1-carbobenzoxy-L- proline (the oil from Part A.) in 175 ml. ofdistilled acetonitrile cooled to there was added 3.46 ml. oftriethylamine followed by 3.34 ml. of isobutylchloroformate. The mixturewas stirred at 0 C. (23) for 15 min. A solution of,6.2 g. of methyla-thiolincosaminide (MTL) from Part B in 85 ml. of water was added, andthe reaction mixture was stirred at 0 C. for 0.5 hr. and at 25 C. for 1hr. The reaction product was then filtered and dried yielding 4.57 g.(37.7%) of methyl N-(4-hutyl- 1-carbobenzoxy L-prolyl) athiolincosaminide. The mother liquor was concentrated under vacuum andan additional 4.25 g. (35.2%) of product recovered. Recrystallizationfrom acetonitrile produced crystals of methyl N-(4-butyl-1-carbobenzoxy-L-prolyl) a thiolincosaminide melting at 194-196C. A second recrystallization from acetonitrile afforded the analyticalsample, M.P. 197.5- 200 C., [d] +111 (c.=0.98, MeOH).

Analysis.-Calcd. for C N N O S: C, 57.75; H, 7.46; N, 5.13; S, 5.93.Found: C, 57.58; H, 7.16; N, 5.50; S, 6.07.

E. METHYL 'N-(4-BUTYLL PROLYL)-a-THIOLLN- COSAMINIDE HYDROCHLORIDE (LE9C4119 A solution of 7.8 g. of methylN-(4-butyl-1-cabobenzoxy-L-prolyl)-a-thiolincosaminide from Part D in200 ml. of methanol was shaken over 2 g. of 10% palladium on carbonunder 40 lbs. of hydrogen pressure for 17 hours. The catalyst wasremoved by filtration and the solution concentrated under vacuum. Theresidue was dissolved in a mixture of 20 ml. of acetone and 20 ml. ofwater and acidified with 6 N hydrochloric acid. Dilution with 4 volumesof acetone precipitated methyl N-(4-butyl-L-prolyl)-ct-thiolincosaminide hydrochloride which was collected byfiltration and dried. The crystals, dried at 55 under vacuum, weighed4.7 g. and melted at l88-194 C. The analytical sample obtained byrecrystallization from acetone melted at 197-199 C. and gave [u] +l5O(water, c.=0.89).

Analysis.-Calcd. for C H N O S-HCh C, 48.80; H, 7.96; N, 6.32; S, 7.24.Found (corrected for 5.54% water): C, 48.58; H, 8.19; N, 6.04; S, 7.36.

This material processes 8% of the antibacterial activity of lincomycinby S. Zuzea assay.

F1. METHYL N- (4-BUTYL-1J IETHYL-L-PROLYL) -a- THIOLINCOSAMINIDE r r na,

N .1101 N .1101 l I 1, COMTL comm 14 acetate, acetone, water (8:4:1) forelution and KMnO; solution for detection consisted chiefly of twomaterials, the cis and trans epimers of methyl N-(4-butyl-1-methyl-L-prolyl)-a-thiolincosaminide in a ratio of about 3 to 2. F2. SEPARATIONOF THE CIS AND TRANS FORMS BY CHROMATOGRAPHY The methyl N (4propyl-1-methyl-L-prolyl)-rx-thi0- lincosaminide from Part F1 wasdissolved in a mixture of methanol and methylene chloride (1:1) and 1.5ml. of triethylamine added. To this solution was added 7 g. of silicagel and the solvent evaporated under vacuum leaving the antibioticdeposited on the silica gel which was sifted on top of a chromatographiccolumn of 200 grams of silica gel packed with a solvent mixtureconsisting of ethyl acetate, acetone, water in a ratio of 8:4:1. Thecolumn was developed by eluting with the same solvent and 20 ml.portions were collected. TLC of each fraction as described above showedthat fractions 31-38, 310 mg, were essentially pure trans epimer andthat fractions 49-74, 32 mg., were essentially pure cis epimer.Fractions 39-48 consisted of a mixture of epimers which could be furtherseparated by repeated chromatography. Each epimer was dissolved in a fewdrops of dilute hydrochloric acid and the hydrochloride precipitated byad dition of acetone. In this manner, there was obtained 50 mg. ofmethyl N (trans 4-butyl-1methyl-L-prolyl)-athiolincosaminide, M.P. -137,and about mg. of methyl N (cis4-butyl-1-methyl-L-prolyl)-a-thiolincosaminide, softening at 105 C. withfurther melting at -185 C.

The trans epimer recrystallized from the same solvent melted at 139-141C. and had the following analysis:

Analysis.Calcd. for C H N O SHCl: Q, H, 8.16; N, 6.13; S, 7.02. Found(corrected for 4.07% H O): C, 48.81; H, 8.54; N, 6.49; S, 6.67.

Similarly recrystallization of the cis epimer gave a product softeningat 108 C. and further at about 189 C. (solvated) with the followinganalysis:

Analysis-Found (corrected for 4.95% water): C, 50.27; H, 9.00; N, 6.05;S, 6.65.

The trans epimer was about 2.2 times as active as lincomycin by S. luteaassay, about 2 times as active by the broth dilution assay, and 2.5times as active in mice infected with S. aureus.

The cis epimer was about /2 to /3 as active as the trans epimer, beingabout equal to lincomycin.

G1. METHYL N-(4-BUTYL-1-ETHYL-L-PROLYL)-ct- THIOLINCOSAMINIDEHYDROCHLORIDE A mixture of 2.0 g. of methyl N-(4-butyl-L-prolyl)-a-thiolincosaminide, 1.5 ml. of acetaldehyde, 150 mg. of 10% palladiumon carbon in 150 ml. of methanol was shaken under 35 lbs. of hydrogenpressure for 5.5 hrs. The catalyst was removed by filtration to give aresidue consisting chiefly of the cis and trans epimers of methyl N-(4-butyl-1-ethyll-prolyl) -u-thi0liI1COSElI11lI'lid6.

G2. SEPARATION OF EPIMERS As described in Part F2, the mixture ofepimers of Part G1, (2 g.) was chromatographed over 200 g. of silica gelusing for elution a solvent system of ethyl acetate, acetone, water(8:4:1), Fractions 33-42 by TLC were pure trans epimer and werecombined, fractions 49-64 were essentially pure cis-epimer and were alsocombined. Fractions 43-48 were a mixture of the epimers which could bepurified by rechromatography. Each epimer was dissolved in a few dropsof dilute hydrochloric acid and the crystalline hydrochloricprecipitated on dilution with a large volume of ether.

The crude trans-epimer fraction of 415 mg. gave 340 mg. (15.4%) ofcrystalline methyl N-(trans-4-butyl-1-ethyl-L-prolyl)-a-thiolineosaminide, M.P. 144-151 C. Recrystallizationfrom dilute acetone raised the M.P. to 148-151" C.

The cis-epimer fraction of 645 mg. afforded 300 mg.

15 (14.1%) of crystalline methyl N-(cis-4-butyl-1-ethyl-L-prolyl)-a-thiolincosaminide, MP. 135l39 C. Recrystallization from diluteacetone gave crystals, M.P. 134- 138 C.

The trans epimer isomer showed about 11.2 times the activity oflincomycin by the S. lutea assay, 24 times the activity of lincomycinagainst Gram positive organisms, and 8 times or more the activity oflincomycin against gram negative organisms. In mice against S. auretzsthe trans epimer was about 2 times as active as lincomycin. The cisepimer was about /2 as active as the trans epimer.

By substituting the butyltriphenyl phosphonium bromide of Part A in theabove example by other substistituent is methyl, ethyl, propyl, pentyl,hexyl, heptyl, octyl, nonyl, decyl, undecyl, and dodecyl, and theisomeric forms thereof, cyclopyropyl, cyclobutyl, cyclopentyl,cyclohexyl, cycloheptyl, cyclooctyl, 2-cyclopropylethyl,3-cyclohexylpropyl, benzyl, phenethyl, 3-phenylpropyl, andtx-naphthylmethyl, the corresponding methyl N-(cisand trans-4-alkyl-,4-cycloalkyl-, and 4 aralkyl 1 carbobenzoxy L prolyl) athiolincosaminides, the corresponding methyl N-(cisand trans-4 alkyl-,4-cycloalkyl-, and 4-aralkyl-L-prolyl)-u-thiolincosaminides; thecorresponding methyl N-(cisand trans-4- 'alkyl-, 4-cycloalkyl-, and4-aralkyl-l-rnethyl-bpropyl)- a-thiolincosaminides and the correspondingmethyl N-(cisand trans-4-alkyl-, 4-cycloalkyl-, and 4aralkyl-1-ethylL-prolyl)-a-thiolincosaminides are obtained. For example, whenpropyltriphenylphosphoniurn bromide and formalin are used lincomycin andallolincomycin (the cisepimer) are obtained both of which are activeantibiotics. By substituting the formalin or acetaldehyde by otherketaldones of the formula R R CO, for example, propionaldehyde, acetone,butyraldehyde, isobutyl methyl ketone, benzaldehyde, phenylacetaldehyde,hydrocinnamaldehyde, acetophenone, propiophenone, butyrophenone, 3methyl 4 phenyl 2 butanone, 2 methyl 5- phenyl-3-pentanone,3-cyclopentanepropionaldehyde, cyclohexaneacetaldehyde,cycloheptanecarboxaldehyde, 2,2- dimethylcyclopropaneacetaldehyde, 2,2dimethylcyclopropyl methyl ketone, cyclopentyl methyl ketone, cyclobutylmethyl ketone, cyclobutanone, cyclohexanone, and 4-methylcyclohexanone,the corresponding methyl N-(cisand trans-4-alkyl-, 4-cycloalkyl-, and4-aralkyl-1-R R CH- L-prolyl)-a-thiolincosaminides where R R CH ispropyl, isopropyl, butyl, 4-methyl-2-pentyl, benzyl, phenethyl,3-phenylpropyl, l-phenylethyl, l-phenylpropyl, l-phenylbutyl,3-methyl-4-phenyl-2-butyl, 2-methyl-5- phenyl-3-pentyl,3-cyclopentylpropyl, 2-cyclohexylethyl, cycloheptylmethyl, 2 (2,2dimethylcyclopropyl)ethyl, 1 (2,2 dimethylcyclopropyl)ethyl,l-cyclopentylethyl, l-cyclobutylethyl, cyclobutyl, cyclohexyl, and4-methylcyclohexyl are obtained. For example, whenethyltriphenylphosphonium bromide and acetaldehyde are used, theantibiotically active methyl 6-(trans-4-ethyl-L-prolylamino) 6,8 dideoxy1 thio D erythro a D- galacto-octopyranoside, methyl6-(trans-4-ethyl-l-ethyl- L prolylarnino) 6,8 dideoxy 1 thio D erythro-D-wgalacto-octopyranoside and the allo forms thereof are obtained.

By substituting the methyl a-thiolincosaminide by otherot-thiolincosaminides or by fl-thiolincosaminides or generally by other6,8-dideoxy-6-amino-D-erythroand L-threo-D-galacto-octypyranosecompounds of Formula 111, examples of which are given hereinafter, thecorresponding amides of Formulas IV and V are obtained.

Example 2.Preparazion of allolincomycin A.4-PROPYLIDENELCARBOBENZOXY-L-PROLINE Sodium hydride (3.8 g.) was warmedwith 75 ml. of dimethylsulfoxide at a temperature of 7075 C. until thereaction Was complete, After cooling to 20 C., 30.8 g.

"tuted 'triphenylphosphonium bromides :where the sub.-.

16 of propyltriphenylphosphonium bromide was added, and the resultingred solution was stirred for 30 minutes to insure complete reaction. Asolution of 5.2 g. of 4-ketol-carbobenzoxy-L-proline in 15 ml. ofdimethyl sulfoxide was added over a period of 15 minutes, and theresulting mixture was stirred for 20 minutes at 26 C. and then at 70 C.for 4 hours. The reaction mixture was cooled, 100 ml. of 5% aqueouspotassium bicarbonate and 100 ml. of water added, and filtered. Thefiltrate was washed twice with 150 ml. portions of ether and the etherwas discarded after back extracting with bicarbonate. The bicarbonatesolutions were combined, diluted with 200 ml. of water, and acidifiedwith 4 N hydrochloricacidlhe acidified" mixture was extracted 7 withthree 200-ml. portions of ether. The combined ether extracts were washedwith three -ml. portions of saturated aqueous sodium bisulfide, thenwith water and dried over anhydrous sodium sulfate. Evaporation of thesolvent gave 5.7 g. of a solid residue which was4-propylidene-l-carbobenzoxy-L-proline.

This residue was dissolved in 18 ml. of acetonitrile and treated with2.8 ml. of dicyclohexylamine. The crystalline dicyclohexylamine salt,5.2 g. yield), melted at 154l57 C. After three recrystallizations fromacetonitrile, an analytical sample was obtained which melted at 164166C. and had a rotation of [a] -8 (c.=0.3898,-CHCl Analysis.-Calcd. for CH N O C, 71.45; H, 9.00; N, 5.95. Found: C, 71.77; H, 9.39; N. 5.1.

Eight grams (17 mmoles) of the dicyclohexylamine salt of4-propylidene-1-carbobenzoxy-L-proline was shaken with excess 1.5 Nsodium hydroxide solution and ether until solution was complete. Thelayers were separated and each one was backwashed. The aqueous alkalinelayer was combined with the backwash from the ether layer and acidifiedwith 4 N hydrochloric acid. The mixture was extracted with ether and theether extracts were combined and evaporated to give 4.8 g. (97.8%) of4-propylidene-l-carbobenzoxy-L-proline as an oil.

l l N MTL COOI-I CsHs 0 GMT L Cal-I To a solution of 2.25 g. of4-propylidene-l-carbobenzoxy-L-pyroline from Part A and 1.40 ml. oftriethylamine in ml. of distilled acetonitrile cooled to 0 there wasadded 1.08 ml. of isobutylchloroformate in 1 ml. of acetonitrile. Themixture was stirred at 0 (15) for 15 min. A solution of 2.92 g. ofmethyl wthiolincosaminide (MTL) in ml. of water was added rapidly. Theresulting solution was stirred at 0 for 1 hr., the cooling bath removedand stirring continued for another hour. The acetonitrile was removed bydistillation under vacuum leaving a partially crystalline residue. Themixture was cooled to 10 and filtered and dried at 55 under vacuum toyield 2.3 g. of crystalline methyl N-(4-propylidene l carbobenzoxyL-prolyl)-wthiolincosamide, M.P. 178186 C. Two recrystallizations frommoist ethyl acetate afforded the analytical sample, M.P. 180187 C., [11]137 (MeOH, c.=0.92).

Analysi.r.-Calcd. for C 'H N O S: C, 57.23; H, 6.92; N, 5.32, S. 6.11.Found: C, 57.24; H, 7.22; N, 5.18;

C. METHYL N- (4-PROPYL-L-PROLYL) -a-THIO- LINCOSAMINIDE HYDROCHLORIDE tt N K \H COMTL COMTL CaHa g A solution of 100 mg. of methylN-(4-propylidene-lcarbobenzoxy-L-prolyl)-a-thiolincosaminide from Part Cin 50 ml. of methanol was shaken over 100 mg. of 7% platinum on Dowex-lunder 40 lbs. of hydrogen pressure for 3 hours. There was then added 100mg. of 10% palladium on charcoal and the reaction mixture was shakenunder 40 lbs. pressure for another 3 hours. The catalyst was removed byfiltration and the solution concentrated under vacuum. The residue wasdissolved in 0.1 m1. of 0.5 N hydrochloric acid. Dilution with 15.0 ml.of acetone precipitated methyl N-(4-propyl-L-prolyD-athiolincosaminidehydrochloride which was collected by filtration. The crystals, dried at55 C. under vacuum weighed 20 mg. and melted at 181-188" C. The productcontained about 4 parts of the cis isomer for each part of the transisomer.

D. METHYL tN-(4-PROPYL-l-METHYL-L-PROLYL)-u.-

THIOLI'NCOSAMINIDE N N Q K J C OMT L C MT L f C H 3 1 One gram of methylN-(4-propyl-l-methyl-L-prolyl)-mthiolincosaminide hydrochloride fromPart D is dissolved in 15-2O ml. of methylene chloride containing 0.5ml. of triethylamine and 2 g. of silica gel for chromatography added.The solvent was evaporated under vacuum leaving the antibiotic depositedon the free flowing silica gel. One hundred grams of silica gel isslurried with 80% aqueous acetone and poured in a chromatographiccolumn. The solvent is drained down to the level of the gel. The sampleof compound and gel prepared previously is dusted on top of the columnfollowed by a layer of sand. The column is eluated with 80% aqueousacetone and fractions of 20 ml. are collected. Each sample is evaporatedand checked by TLC on silica gel using 80% aqueous acetone as theeluting solvent. The antibiotic is detected by spraying with alkalinepermanganate solution. The fractions containing the desired products arecombined and crystallized as the hydrochloride by dissolving in excessdilute hydrochloric acid and diluting with acetone. The trans isomer iseluated first followed by a mixture of cis and trans isomers and finallypure cis isomer. The mixture may be rechromatographed as above. The cisisomer, allolincomycin, had a melting point of 147- 150' C., an [ab 110(H 0), and the following elemental analysis:

Calculated for C gH N O S'HCl2 C, 48.80; H, 7.96; N, 6.32. Found(corrected for 9.47% H O): C, 49.15; H, 7.80; N, 6.39.

By substituting the propyltriphenylphosphonium bromide of Part A in theabove example by other substituted triphenylphosphonium bromides wherethe substituent is butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl,undecyl, and dodecyl and the isomeric forms thereof as well as methyl,ethyl, and isopropyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl,cycloheptyl, cyclooctyl, 2- cyclopropylethyl, 3-cyclohexylpropyl,benzyl, phenethyl, 3-phenylpropyl, and u-naphthylmethyl, thecorresponding methyl N-(cis and trans-4-alkylidene, cycloalkylidene-,and aralkylidene 1-carbobenzoxy-L-prolyl)-a-thiolincosaminides, thecorresponding methyl N-(cis and trans-4- alkyl, cycloalkyl-, andaralkyl-L-prolyl)-a-thiolincosaminide, and the corresponding N-(cis andtrans-4-alkyl-, cycloalkyl-, and .aralkyl 1methyl-L-prolyD-a-thiolincosaminides are obtained. By substituting theformalin of Part D by other ketaldones of the formula R R CO, forexample, acetaldehyde, propionaldehyde, acetone, butyraldehyde, isobutylmethyl ketone (4-methyl-2-pentanone), benzaldehyde, phenylacetaldehyde,hydrocinnamaldehyde, aeetophenone, propiophenone, butyrophenone, 3methyl-4-phenyl-Z-butanone, 2-methyl-5-phenyl-3-pentanone, 3cyclopentanepropionaldehyde, cyclohexaneacetaldehyde,cycloheptanecarboxaldehyde, 2,2-dimethylcyclopropaneacetaldehyde,2,2-dimethylcyclopropyl methyl ketone, cyclopentyl methyl ketone,cyclobutyl methyl ketone, cyclobutanone, cyclohexanone, and4-methylcyclo hexanone, the corresponding methyl N-(cis andtrans-4-alkyl-, 4-cycloalkyl-, and 4-aralkyl-1-R R CH-L-prolyl)-a-thiolincosaminides where R R CH is ethyl, propyl, isopropyl,butyl, 4-methyl-2-pentyl, benzyl, phenethyl, 3-phenylpropyl,l-phenylethyl, l-phenylpropyl, 1- phenylbutyl,3-methyl-4-phenyl-2-butyl, 2-methyl-5-phenyl-3-pentyl,3-cyclopentyl-propyl, 2-cyclohexylethyl, cycloheptylmethyl,2(2,2-dimethylcyclopropyl)ethyl, 1-(2, Z-dimethylcyclopropyl)ethyl,l-cyclopentylethyl, l-cyclobutylet'hyl, cyclobutyl, cyclohexyl, and4-rnethylcyclohex- 311 are obtained.

By substituting the methyl a-thiolincosarninide by othera-thiolincosaminides or by fl-thiolincosaminides or generally by other6,8-dideoxy-6-amino-D-erythro-D-galactooctopyranose compounds of FormulaIII, examples of which are given hereinafter, the corresponding amidesof Formulas IV and V are obtained.

Example 3.-Ethyl u-thz'olz'ncosaminide Lincomycin C hydrochloride (2 g.)was dissolved in 50 ml. of water. The pH of the solution was adjusted to9.5 by the addition of an anion exchange resin in the hydroxide form.(An anion exchange resin obtained by chlorometylating by the proceduregiven on pages 88 and 97 of Kunin, Ion Exchange Resins, 2nd ed., [1958],John Wiley and Sons, Inc., polystyrene cross-linked, if desired, withdivinylbenzene, prepared by the procedure given on page 84 of Kunin,supra, and quaternizing with trimethylamine or dirnethylethanolamine, bythe procedure given on page 97 of Kunin, supra.) The alkaline solutionwas then freeze dried to a residue which was dissolved in 50 ml. ofhydrazine hydrate (98100%) and refluxed for 24 hours. The solution wasthen concentrated to dryness in vacuo and the residue triturated threetimes with 10 ml. portions of acetonitrile. The insoluble material wascollected and dried; yield 900 mg. A solution of 600 mg. of the driedinsoluble material in 4 m1. of dimethylformamide (heat was used topromote solution) was then clarified by filtration and the filtrate washeld at room temperature for 4 hours. The crystalline ethyla-tl'liOllIlCOS aminide which precipitated was isolated by filtration,washed with ether and dried; yield 500 mg.

Ethyl u-thiolincosaminide has the following physical and chemicalcharacteristics: Melting Point: 191-195 C.; Optical Rotation: [a] +258(c., 0.76 in water); and Titration: pKa=7.17, and the followingelemental analysis:

Calculated for C H NO S: C, 44.93; H, 7.92; N, 5.24; S, 11.99; 0, 29.92.Found: C, 44.09; H, 7.91; N, 5.24; S, 11.32.

The lincomycin C hydrochloride was prepared asfollows:

' FERMENTATION A soil slant of Streptomyces lincolnensis var.lincolnensis, NRRL 2936, was used to inoculate a series of 500- ml.Erlenmeyer flasks each containing 100 ml. of seed medium consisting ofthe following ingredients:

Yeastolac 1 g 10 Glucose monohydrate g 10 N-Z-amine B 2 g 5 Tap waterq.s. liters 1 Yeastolac is a protein hydrolysate of yeast cells.

'-N-Z-amine B is Shelfields enzymatic digest casein.

The seed medium presterilization pH was 7.3. The seed was grown for 2days at 28 C. on a Gump rotary shaker operating at 250 r.p.m.

A 5% inoculum of the seed described above (5 ml.) was added to each of30 500-ml. Erlenmeyer flasks each p 1 Wilsons Peptone Liquor No. 150 isa preparation of enzymatically hydrolyzed proteins from animal origin.

At the time of inoculation, DL-ethionine was added to a finalconcentration of 2 mg./ml.

The shake flasks were harvested after 4 days of fermentation at 28 C. ona Gump rotary shaker at 250 r.p.m. They assayed 200 meg/ml. on the S.lutea assay, hereinafter described. The whole beer solids was about 20gin/liter.

PURIFICATION Whole beer (235 liters) from a DL-ethionine fermentationwas filtered at harvest pH using a filter aid as required. The mycelialcake was washed with water and the cake was then discarded. The filteredbeer and water wash (275 liters) was stirred for minutes with 12.5 kg.of activated carbon and 2.5 kg. of diatomaceous earth. The mixture wasfiltered and the filtrate was discarded. The carbon cake was washed withliters of water and the water wash was discarded. The cake was washedwith liters of 20% aqueous acetone and the 20% aqueous acetone wash wasdiscarded. The cake was then eluted twice with 100 liter portions ofaqueous acetone. The eluates were combined (215 liters) and the solutionwas concentrated (18 liters). This concentrate was adjusted to pH 10.0with a 50% aqueous sodium hydroxide solution and extracted three timeswith 20 liter portions of methylene chloride. The methylene chlorideextracts were combined (60 liters) and then concentrated to give an oilypreparation (7.14 g.) containing lincomycin and lincomycin C in equalamounts and both in the free base form. This preparation was thendissolved in 200 ml. of methylene chloride. The solution was clarifiedby filtration and then concentrated to dryness in vacuo. The residue wasdissolved in ml. of 1 N methanolic hydrogen chloride. The methanolicsolution was then mixed with 3.2 liters of ether under stirring. Theresulting precipitated colorless, crude lincomycin hydrochloride andlincomycin C hydrochloride was isolated by filtration and dried; yield7.14 g. assaying 940 mcg./ mg. against Sarcina lutea. (The assay againstSarcina lutea is conducted on agar buffered to pH 6-8 with pH 7.0phosphate buffer [0.1M]. A unit volume [0.08 ml.] of solution containingthe material to be assayed is placed on a 12.7 ml. assay disc which isthen placed on an agar plate seeded with the assay microorganism.) Thinlayer chromatography showed the presence of both lincomycinhydrochloride and lincomycin C hydrochloride in approximately equalamounts.

Crude lincomycin C hydrochloride (7.0 g.) was dissolved in 20 ml. ofwater and 20 ml. of butanol, pH adjusted to 4.2 with 1N HCl, and thesolution distributed in a counter current distribution apparatus for1,000 transfers. Analysis by thin-layer chromatography showed that thefractions in tubes to 190 contained lincomycin C. These fractions werecombined, and the solution was concentrated to an aqueous and freezedried to give 2.44 g. of lincomycin C hydrochloride assaying 1400meg/mg. against Sarcina lutea. Five hundred mg. of this preparation wasdissolved in 2 ml. of water, 1 ml. of methanol, and 100 ml. of acetone.The solution was clarified by filtration. The filtrate was mixed withether until crystals appeared. The mixture was allowed to stand at roomtemperature for 1 hr. Crystalline (cubes) lincomycin C hydrochloride wasseparated from the supernatant material solution by decantation. Thesecrystals were recrystallized from one ml. of water, one ml. of methanol,80' ml. of acetone and 20 ml. of ether; yield, 250 mg. of crystalline(cubes) lincomycin C hydrochloride. The supernatant (obtained asdescribed above) was allowed to stand at 5 C. for 4 hours. Crystalline(needles) lincomycin C hydrochloride which precipitated was filtered anddried; yield, mg. of crystalline (needles) lincomycin C hydrochloride.

Following the procedure of Example 1, compounds of Formulas IV and Vwherein R, R and Z are as given above and Y is -SCH CI-I in thea-configuration are obtained. For example, when ethyl orpropyltriphenylphosphonium bromide and formalin or acetaldehyde are usedantibiotically active ethyl 6-(trans-4-ethyl-L-prolylamino)-, ethyl6-(trans-4-ethyl-1-methyl-L-prolyl)-, ethyl 6-trans-4-ethyl-1-ethyl-L-prolyl) ethyl 6-(trans-4-propyl-L-prolyl)-,ethyl 6-(trans-4-propyl-l-rnethyl-L-prolyl),- and ethyl6-(trans-4-propyl-l-ethyl-L-prolyl-6,8-dideoxy1-thio-D-erythro-ot-D-galacto-octopyranosides and the allo forms thereofare obtained.

Example 4.Alkyl fl-thiolincosaminides A. METHYLN-ACETYL-a-THIOLINCOSAMI'NIDE Five grams of methyl ot-thiolincosaminide(about 0.02 mole) was suspended with stirring in 50 ml. of methanol andtreated with 4.04 g. (about 0.04 mole) of acetic anhydride. The startingmaterial dissolved almost completely, and thereafter the mixture becamesolid. After 18 hours at room temperature (about 25 C.), the solid wasfiltered, washed with methanol, and dried in a vacuum oven at 50 C. and15 mm. pressure; yield, 4.58 g. (79%) of crystalline product melting at242-245 C. This product was recrystallized from absolute methanol togive colorless needles of methyl N-acetyl-a-thiolincosaminide having amelting point of 243-245 C. and a rotation of [M +265 (c., 0.7374,water).

Analysis.-Calcd. for C H NO S: C, 44.72; H, 7.17; N, 4.74; S, 10.85.Found: C, 44.87; H, 7.10; N, 4.65; S, 10.99.

B. MIXTURE OF G-ACETAMIDO-6,8-DIDEOXY-D ERYTH- RO-a- ANDB-DGALACTO-OCTOPYRANOSES(N-ACETYL- a and fi-LINCOSAMINES) A solution of50 g. of methyl N acetyl-a-thiolincosaminide of part A in 1500 ml. ofwater at 40 C. was stirred magnetically and treated with a solution of70 g. of mercuric chloride in 1500 ml. of water at 40 C.; a whiteprecipitate for-med immediately. The reaction was continued withoccasional heating to 40 C. for a period of 3 days. Thin-layerchromatography then showed the absence of starting material. Theprecipitate of mercuric chloride mercaptide (ClHgSMe) was removed byfiltration. The colorless filtrate, together with aqueous washes of theprecipitate, was stirred magnetically at room temperature, and theexcess of mercuric chloride was removed by the addition of pyridine insmall portions until the precipitation of the insoluble mercuricchloride-pyridine complex was complete. After storing the mixture in therefrigerator at C. for 3 hours, the solid was removed by filtration, theprecipitate was washed well with cold water, and the combined filtrateand washings were stirred with a small amount of silver carbonate untilthe solution was neutral to pH paper. The solution was thereuponfiltered through a Millipore filter (Millipore Filter Corporation,Bedford, Mass), the filter was washed thoroughly with water, and thewashings were added to the colorless filtrate. Excess silver ion wasprecipitated by saturating the solution with hydrogen sulfide andremoving the silver sulfide by filtration. The silver sulfide was washedwith water, the washings were added to the filtrate, and the solutionwas lyophilized to give a colorless amorphous solid, which was a mixtureof 6-acetamido-6,8-dideoxy- D-erythro-aand ,B-D-galacto-octopyranoses(N-acetyl-ixlincosamine and N-acetyl-fl-lincosamine).

C. N -A CETYL-LZ, 3,4,7PENTAOA'CETYL-B-LI'NCOSAMI'NE The crude mixtureof N-acetyl-u-lincosamine and N- acetyl-fi-lincosamine of part B wasslurried in 400 ml. of pyridine and 200 ml. of acetic anhydride andstirred magnetically overnight at room temperature. The resultingcolorless solution was concentrated on a rotating evaporator at 40 C.and 1 mm. Hg pressure to a pale yellow syrup which was dissolved in amixture of water and chloroform. The aqueous layer was extracted withchloroform and the chloroform extracts were combined, washed with dilutesulfuric acid (2 N), then twice with water, with saturated aqueoussodium bicarbonate, with water until neutral, and finally dried overanhydrous sodium sulfate. The resulting chloroform extract was thenevaporated in a rotating evaporator at 35 C. and ml. Hg pressure to givea colorless solid which was dissolved in hot ethyl acetate. To thisethyl acetate solution was added Skellysolve B hexanes until solidsbegan to crystallize. The solids were removed by filtration and twicerecrystallized from ethyl acetate-Skellysolve B hexanes to give N-acetyl-1,2,3,4,7-penta-O-acetyl-,B-lincosamine (9.27 g.) of meltingpoint 227-230 C. and rotation +33 (c., 0.832, chloroform).

Analysis.-Calcd. for C H NO C, 50.52; H, 6.15; N, 2.95. Found: C, 50.40;H, 6.42; N, 3.04.

The ethyl acetate-Skellysolve B hexanes filtrates from the firstcrystallization were allowed to stand at room temperature, whereuponcolorless prismatic needles of-acetyl-1,2,3,4,7-penta-O-acetyl-wlincosamine separated; M.P. 169172 C.The melt resolidified on cooling in the form of hexagonal plateletswhich then melted at 237- 238 C.; after recrystallization from ethylacetate-Skellysolve B hexagonal platelets were obtained melting at 240-240.5 C. and having a rotation of [a] +132 (c., 0.9842, chloroform).

Analysis.-Calcd. for C H NO C, 50.52; H, 6.15; N, 2.95. Found: C, 50.62;H, 6.08; N, 3.02.

The N acetyl 1,2,3,4,7 penta O acetyl a and B- lincosamines can also beseparated by counter current distribution, using a system ofwater:acetone:methyl ethyl ketone:cyclohexane in a volume ratio of3:5:4:4. In 900 transfers, approximately 50% of each of the 2 anomerscould be isolated in the pure state, the partition coefiicients beingfor the a-anomer, 0.75; for the ,B-anomer, 0.66.

22 D. G-ACETYLAMINO 1a BROMO 1,6,8 TRIDEOXY-D- ERYTHRO DGALACTO-OCTOPYRA NOSE ('N-ACETYL- 2,3,4,7-TETRA-O-ACETYL-1a-BROMO 1DEOXYLINCOS- AMI-NE) OAc XXXVI Two grams ofN-acetyl-1,2,3,4,7-penta-O-acetyl-fl lincosamine of Part C were stirredmagnetically with a solution of anhydrous hydrogen bromide in aceticacid in 5 ml. of acetic acid (saturated at 0) at room temperature (about25 C.) for about 3 hours. All of the solids dissolved within one hour.The pale yellow, viscous solution was diluted with 50 ml. of chloroform,poured onto ice and stirred for 10 minutes. The chloroform layer wasseparated, the aqueous solution was extracted thoroughly withchloroform, and the combined chloroform extracts were washed with wateruntil the aqueous wash was new tral to Congo red paper, and dried overanhydrous sodium sulfate. The chloroform was thereupon evaporated in arotatory evaporator at C. and 15 mm. Hg pressure to give an almostcolorless amorphous solid. After three crystallizations of the solidfrom chloroform-Skellysolve B, colorless, prismatic needles ofN-acetyl-2,3,4,7-tetra-O- acetyl-1u-bromo-l-deoxylincosamine wereobtained which had a melting point of 188189 C. and a rotation of [a]+23l (c., 0.8132, chloroform).

Analysis.-Calcd. for C H BrNO C, 43.56; H, 5.28; N, 2.82; Br, 16.10.Found: C, 43.68; H, 5.39; N, 2.88; Br, 17.22.

E. METHYL rN-ACETYL-2,3,4,7-TETRA-O-ACETYL-fl- THIOLI-N'COSAMINIDE Twograms of N-acetyl-2,3,4,7-tetra-O-acetyl-lot-bromol-deoxylincosamine ofPart D were dissolved in 25 ml. of acetone which had been driedpreviously over potassium carbonate, and to this solution were added 350mg. of thiourea. A-fter warming briefly on a steam bath until the solidshad dissolved, the colorless reaction mixture was left overnight at roomtemperature. To it was then added a solution of 680 mg. of potassiumcarbonate and 860 mg. of sodium bisulfite in 10 ml. of water, followedby 900 mg. (0.40 ml.) of methyl iodide. The mixture was kept in astoppcred bottle which was shaken mechanically at room temperature for 3hours. The reaction mixture was then extracted thoroughly withchloroform, the combined extracts were washed twice with water, driedover anhydrous sodium sulfate, and the solvents were removed on arotating evaporator at 40 C. and 15 mm. Hg pressure. In this manner 1.38g. of colorless, amorphous solid were obtained, which upon thin-layerchromatography was shown to be a single compound. This solid wasdissolved in hot ethyl acetate which was thereupon diluted withSkellysolve B hexanes to give 880 mg, of colorless platelets of meltingpoint 268272 C. Recrystallization from the same solvents gave methylN-acetyl-Z,3,4,7-tetra-O-acetyl- ,B-thiolincosaminide of melting point272273 C. and rotation [0c] I-3l (0., 0.6800, chloroform).

Analysis.Calcd. for C H NO S: C, 49.22; H, 6.31; N, 3.02; S, 6.92.Found: C, 49.15; H, 6.23; N, 3.00; S, 6.41.

F. METHYL :N-ACETYL-fi-THI OLINCOS-AMI NIDE Ten ml. of methanol weresaturated with dry ammonia gas at a temperature between 0 and 5 C. Intothis saturated solution was added at room temperature 1 g. of methyl Nacetyl 2,3,4,7 tetra O acetyl 3 thiolincosaminide. The mixture wasallowed to stand at room temperature for three hours and then taken todryness on 23 a rotating evaporator at 40 C. and mm. Hg pressure, andthe solid thus obtained was recrystallized three times from ethanol togive methyl N-acetyl-fi-thiolincosaminide.

G. MTETHYL fi-THI OLI N-C OSAMINIDE A solution of 1 g. of methylN-acetyl-B-thiolincosaminide in 10 ml. of hydrazine hydrate was refluxedfor 24 hours. Excess hydrazine was then removed in a stream of drynitrogen on a steam bath. The crystalline residue was recrystallizedfrom 5 ml. of water, the crystals were collected, washed with coldwater, and dried in vacuo to give methyl p-thiolincosaminide.

By substituting the methyl iodide of Part E by ethyl,

propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl,.decyl,

undecyl, and dodecyl iodides and the isomeric forms thereof thecorresponding alkyl N-acetyl-l,2,3,4,7-tetra-O-acetyl-fi-thiolincosaminides, alkyl N-acetyl-B-thiolincosaminides, andalkyl fi-thiolincosaminides are obtained.

Following the procedure of Example 1 compounds of the formula XXX VIIwherein X=R or Z as given above and R is as given above are obtained.

By substituting 2-hydroxyethyl iodide for the methyl iodide of Part E,2-hydroxyethyl N-acetyl-2,3,4,7-tetra-O- acetyl-B-thiolincosaminides,2-hydroxyethyl N acetyl ,8-

thiolincosaminide, and 2-hydroxyethyl-B-thiolincosamina 2-alkoxyethyliodide the corresponding Z-alkoxyethyl compounds are obtained. The alkylof the alkoxy can be methyl, ethyl, propyl, butyl, pentyl, hexyl,heptyl, octyl, nonyl, decyl, undecyl, or dodecyl or the isomeric formsthereof.

Following the procedures of Example 1, compounds of the formula:

ide are obtained. By using A. METHYL N-ACETYL-S,4-O-ISOPROPYLIDE'NE-fl-THI OLI-NC OSAMINIDE A suspenesion of 5.3 g. of finely-powdered methylN-acetyl-B-thiolincosaminide was stirred for minutes at room temperaturewith 500 ml. of acetone and 5 ml. of concentrated sulfuric acid. After30 minutes another 5 ml. of concentrated sulfuric acid was added, andthe solution was stirred for another half hour at room temperature. A

24 suspension of 150 g. of barium carbonate in ml. of water was added,and the mixture was stirred until neutral. The barium sulfate and excesscarbonate were removed by filtration, and the precipitate was washedwith acetone. The filtrate and washings were combined and evaporated todryness in vacuo at 40 C. The residue was treated with acetone:ether(10:1) and insoluble materials were removed by filtration. The filtratewas evaporated to dryness and the residue was dissolved in 100 ml. ofethanol. Removal of the alcohol by heating in vacuo left a gummymaterial which was dissolved in 20 ml. of warm water containing a smallamount of sodium carbonate. After removal of some insoluble material byfiltration, the filtrate was cooled. The. crystals which formed werecollected after standing in the refrigerator for 4 hours,

washed with cold water, and dried in vacuo, to give methylN-acetyl-3,4-O-isopropylidene-fl-thiolincosaminide.

B. lN-ACETYL-3,4-O-ISOPROPYLIDENE-1-DE OXY- LI'NCOSAMIN E Five grams ofmethyl N-acetyl-3,4-O-isopropylidene-flthiolincosaminide was heatedunder reflux for 7 hours with 35 ml. of loosely-packed Raney nickel inml. of ethanol. The mixture was thereupon filtered and the catalyst waswashed with a total of 400 ml. of boiling ethanol. The filtrate andwashings were combined and evaporated to dryness, leaving a partiallycrystalline residue. This residue was purified =by counter currentdistribution in a system of butanolzwater to giveN-acetyl-3,4-O-isopropylidene-l-deoxylinc0samine.

C. I-DEOXYLINCOS AMINE One gram of N-acetyl-3,4-O-isopnopylidene-l-deoxy? lincosamine in 5 ml. of hydrazine hydrate(98-100%) is heated under reflux for 21 hours. The excess hydrazinehydrate is distilled in vacuo. The residue is stirred with acetonitrileand again evaporated. Five ml. of water is added and the pH adjusted to2 with hydrochloric acid. After 3-5 hrs. at 26 the solution is dilutedwith 10 ml. of water and excess silver carbonate added. The mixture isthen filtered and the filtrate lyophilized. The resulting residue iscrystallized from water to yield l-deoxylincosamine.

Following the procedure of Example 1, compounds of the formula:

wherein X==R or Z as given above and R is as given above are obtained.

\' l l/SCHzCIIaOI-I 0 II XLI A. 2-HYDROXYETHYL a-THIOCELESTOSAMI'NIDEHYDRAZINE 'SOLVA-TE A mixture of 5 g. (0.0094 mole) of celesticetin(Example 3, US. Patent 2,928,844) and 25 ml. (excess) of hydrazinehydrate was heated under reflux for 21 hours. The excess hydrazine wasremoved by distillation in vacuo and the residue was crystallized from35 ml. of absolute ethanol. White crystals (1.2 g.) of 2-hydroxyethyla-thiocelestosaminide hydrazine solvate were obtained which melted at98-108 C. Recrystallization lfrom absolute ethanol gave 0.65 g. havingan optical rotation of [a] -=+243 (c.=0.8, Water); an infraredabsorption spectrum in Nujol mull at the following frequencies: 3,400,1,630, 1,600, 1,460, 1,450 (sh), 1,305, 1,275, 1,260, 1,200, 1,115,1,085, 1,055, 1,010, 978, 950, 925, 910, 873, 800-820, 705, 690, and 680cmf an equivalent weight of 161 with two basic groups having pKas in theregion of 7.5; and the following elemental analysis:

Calculated for C H NO S-N H C, 40.11; H, 8.26; N, 12.76; S, 9.74. Found:C, 40.15; H, 8.04; N, 11.69; S, 9.56.

B. 2-HYDROXYETHYL a-THIOCELESTOSAMINIDE HYDRAZINE SOLVATE Desalicetin(Example 1, US. Patent 2,851,463) (10 g.) was dissolved in hydrazinehydrate (100 cc.) and heated under reflux in an oil bath at 165 C. for18 hours. The almost colorless solution was concentrated to dryness on asteam bath, first at mm., and finally at less than 1 mm., giving a solidresidue which was triturated with acetonitrile, filtered, and washedwith the same solvent. Crystallization from ethanol gave colorlessneedles (2.64 g.) of 2-hydroxyethyl a-thiocelestosaminide hydrazinesolvate. Recrystallization from the same solvent gave crystals ofZ-hydroxyethyl a-thiocelestosaminide hydrazine solvate which had anequivalent weight of 168 with two basic groups having pKas in the regionof 7.5; an optical rotation of [a] =-|-Z48 (c.=l, 95% ethanol); and thefollowing elemental analysis:

Calculated for C H NO S-N H N, 12.76; S, 9.74. Found: C, 40.27; S, 9.80.

C, 40.11; H, 8.26; H, 7.95; N, 11.63;

C. Z-HYDR'OXYETHYL d-THIOCELESTO SAMINIDE A solution of 2 g. of2-hydroxyethyl a-thiocelestosaminide hydrazine solvate, prepared as inPart A or Part B, in ml. of dimethylformamide was concentrated to avolume of about 10 ml. and then diluted with 10 ml. ofdimethylformamide. sulted, and 500 mg. nide was deposited. Thecrystalline 2-hydroxyethyl a-thiocelestosaminide had an optical rotationof [a] =|-262 (c.=l, in water); an infrared absorption spectrum at thefollowing frequencies: 3,400 (sh), 3,250, 1,600, 1,400, 1,325, 1,310,1,290, 1,240, 1,160, 1150 (sh), 1,110, 1,100 1,075, 1,045, 1,038, 1,005,980, 920, 895, 862, 825, 796, 740, 711, and 690 cmf an equivalent weightof 297 and a pKa of 7.2; and the following elemental analysis:

Calculated for S H NO S: C, 44.43; H, 7.80; N, 4.71; S, 10.78. Found: C,44.20; H, 7.78; N, 4.97; S, 10.68.

Following the procedure of Example 1, compounds of the formula:

X BIT CH3 orr.o I\ (L-NH- H RH HO K on A \I I/ S-CHzCHzOH OH XLII givenabove and R is as given Ether was added until cloudiness reof2-hydroxyethyl a-thiocelestosami- 0 26 Example7.-7-0-methyl-1-deoxylinc0samine CHaO" HzN- A. 2-HYDROXYETHYL N-ACETYL-3,4-O-ISOPROPYLI- DENE-a-THIOCELESTOSAMINIDE To a solution of 14g. (0.047 mole) of 2-hydroxyethyl u-thiocelestosaminide in 150 ml. ofethanol were added 14 ml. of acetic anhydride. The reaction mixture wasstirred for /2 hour, refrigerated overnight, and evaporated to drynessin vacuo. The residue, 2-hydroxyethyl N-acetyla-thiocelestosaminide, wasslurried with ether and dried under reduced pressure.

The thus-obtained dried material was dissolved in 1,500 ml. of acetoneand 15 ml. of concentrated sulfuric acid were added to the solution withstirring. The mixture was stirred for about 2 hours and then neutralizedby adding dry ammonia. The mixture was filtered and the filtrate wasevaporated to dryness, leaving 2-hydroxyethyl N- acetyl 3,4 Oisopropylidene a thiocelestosaminide as an oil.

B1. rN-ACETYL-3A-O-I SOPR OPYLIDENE-7 -O-METHYL-1- DE OXYLI'NCOSAMIN EThe oil of Part A was dissolved in 5 00 ml. of ethanol, 150 ml. ofloosely-packed Raney nickel in ethanol were added thereto, and themixture was heated under reflux for 10 hours. Thereafter the mixture wasfiltered, the catalyst was washed with 1 liter of boiling ethanol andthe combined filtrate and washings were evaporated to dryness. An oilresulted which was distributed (500 transfers) in the systeml-butanol:water. A peak fraction, K=0.82, obtained by evaporation ofpooled tubes 200- 250, gave 4.6 g. (33%) ofN-acetyl-3,4-O-isopropylidene- 7-O-methyl-l-deoxylincosamine whichcrystallized upon drying; melting point, 198-205 C.; rotation [a] +71(c. 1, 50% ethanol).

Analysis.-Calcd for C H NO C, 55.43; H, 8.31; N, 4.62; Methoxyl, 10.63.Found: C, 55.03; H, 8.28; N, 4.70; Methoxyl, 10.43.

Potassium metal (1.16 g.) was dissolved in ml. of tert-butyl alcohol.The solvent was removed as completely as possible by distillation atatmospheric pressure and finally by vacuum distillation at 15 mm. Hgpressure. To the dry solid residue was added 100 ml. of dry benzene andthe benzene was removed by distillation to leave a fine powder. To thethus-obtained powdery potassium tert-butoxide was added 200 ml. of drybenzene, and the mixture was stirred magnetically at room temperatureuntil an opalescent solution resulted. To this solution was added 5 g.of N-acetyl-3,4,-O-isopropylidene-l-deoxylincosamine, prepared accordingto Part B of Example 5, and the mixture was then stirred over night atroom temperature. To this mixture was added 42.4 g. (18.6 ml.) of methyliodide and the mixture was stirred at room temperature for 2 hours. Themixture was filtered to remove potassium iodide, and the filtrate wasdistilled in vacuum at about 35 C. to give a colorless syrupy material,which was subjected to counter current distribution in a system of ethylacetatezethanokwater in a ratio of 4:1:2. The fractions containingN-acetyl-3,4-O-isopropylidene-7-O- methyl-l-deoxylincosamine, asdetermined by thin-layer chromatography, were combined, evaporated todryness and the residue was crystallized to give pure N-acetyl-3,4-O-isopropylidene-7-O-methyl-l-deoxylincosamine.

C. 7-O-METHYL-1-DEOXYLINCOSAMINE Following the procedure of Example 5,Part C, N- acetyl 3,4 O-isopropylidene-7-O-methyl-l-deoxylincosamine isconverted to 7-O-methyl-l-deoxylincosamine.

Following the procedure of Example 1, compounds of the formula:

Example 8.2-Hydr0xyethyl OL-thlOllilCOSdflliilidE A. METHYL'NACETYL-2,3,4,7-TETRAO-THIO'NO BENZOYL-a-THIOLINCOSAMINIDE MethylN-acetyl-tx-thiolincosaminide (IX), dissolved in dry pyridine, istreated with an excess of thiobenzoyl chloride (180% excess) at C. andthe reaction mixture allowed to stand overnight at room temperature withthe exclusion of atmospheric moisture. Following the removal ascompletely as possible of volatile material at 30 C. under high vacuum,the reaction product is extra-cted with chloroform, the extract washedwith water, dilute sulfuric acid (N), water, saturated aqueous sodiumbicarbonate, with water again, and dried over anhydrous sodium sulfate,Removal of the chloroform in vacuo yields the product, methylN-acetyl-2,3,4,7-tetra-O-thionobenzoyl-a-thiolincosamini-de (X), whichcan be recrystal lized from acetone or ethyl acetate by the addition ofSkellysolve B (technical hexane).

B. Z-HYDROXYETHYLATION or N-ACETYL-Z-O-BEN-ZOYL-3,4,7-TRI-O-THIONOBENZOYL-a-THIOLI'NCOSAMI- \NIDES Thetetrathionobenzoate (X) of part A is dissolved in ethanol-freechloroform (to give a 2 to solution) and treated with a solution ofbromine (2 molecular equivalents) in chloroform (a 12% solution,vol/vol.) with magnetic stirring at room temperature with the exclusionof atmospheric moisture. After 2 hours the volatile material is removedas completely as possible by evaporation at 30 C. and mm. Hg pressure,more chloroform (ca. 100 ml. per 2 g. of starting material) added, andremoved as before. To the residue is then added acetone (ca. ml. per 2g. of starting material) and triethylamine (2 molar equivalents), andthe resulting solution heated under reflux on a steam bath for 1 hr.,with the exclusion of atmospheric moisture. [The initial brominationproduce (XI), is thus converted to the cyclised intermediate (XII).] Tothe cooled solution is then added potassium carbonate (5 molecularequivalents), sodium bicarbonate (wt. equal to the K CO use-d), andwater (ca. 10 ml. per g. of K CO followed by ethylene iodohydrin(preferably, ca. 610 molecular equivalents), and the stoppered mixtureis shaken mechanically for 3 hrs. at room temperature. The reactionmixture is extracted with chloroform followed by washing with water toremove inorganic salts, and drying over anhydrous sodium sulfate.Removal of the solvent at C, in vacuo yields 2-hydroxyethylN-acetyl-2-O-benzoyl- 3,4,7-tri-O-thionobenzoyl-a-thiolincosaminide.

The above acylated compound is heated under reflux with hydrazinehydrate (20-30 times the weight of the ester) for 1236 hours and theexcess hydrazine then removed by distillation under reduced pressure.The residue is triturated with acetonitrile, the solid product removedby filtration, washed with a-cetonitrile and dried by 23recrystallization from ethanol yields 2-hydrox-yethylotthiolincosaminide.

By using a Z-alkoxyethyl iodide the corresponding 2- alkyloxyethylcompounds are obtained. The alkyl of the alkyloxy can be methyl, ethyl,propyl, butyl, pent-yl, hexyl, heptyl, octyl, nonyl, decyl, undecyl, ordodecyl or the isomeric forms thereof.

Following the procedure of Example 1, compounds of the formula: I

g\ofi //S|C.H2CII2OR4 XLV wherein X=R or Z as given above and R and Rare as given above, are obtained.

Example 9.Alkyl ot-thiolincosaminides I-I2N N l/ lii: ii no S-alky 0HXLVII wherein X=R or Z as given above and R is as given above, areobtained.

Example 10.2-O- and 7-0-alkylari0n of alkyl a-and ,B-thiolincosaminidesA. METHYL N-ACETYL3,4O-ISOPROPYLIDENE-a-THIO- LINCOSAMINIDE AND THEOXAZOLI NE DERIVED THEREFROM Sixteen grams of methylN-acetyl-a-thiolincosaminide was finely powdered and suspended in 1,600ml. of dry acetone with rapid magnetic stirring. To this suspension wasadded 16 ml. of concentrated sulfuric acid. The suspended solid began todissolve and solution was complete within 30 to minutes. After standingfor 3 hours at room temperature (2426 C.), the solution was cooledovernight in a refrigerator at about to C.

The pale yellow solution was neutralized by introducing, with stirring,a stream of dry ammonia gas. Ammonium sulfate precipitated and wasremoved by filtration and washed with acetone. The acetone washing wasadded to the colorless filtrate which was evaporated on a rotatingevaporator at 30 C. and mm. Hg pressure to yield a mixture of syrup andcrystalline solids. The syrup was dissolved by swirling the mixture with50 ml. of water, and the crystalline solid was collected on a filter andwashed with ice-cold water. Upon drying at 60 C. and 15 mm. Hg pressure,7.12 g. of solids were obtained of melting point 189192 C.Recrystallization from acetone-Skellysolve B hexanes gave colorlessneedles of the oxazoline derived from methylN-acetyl-3,4-O-isopropylidene-u-thiolincosaminide; melting point,191-192.5 C.; rotation [a] +l26 (c., 0.8508, ethanol).

Analysis.-Calcd. for C H NO S: C, 52.95; H, 7.30; N, 4.41; S, 10.10; 0,25.20. Found: C, 52.77; H, 7.34; N, 4.40; S, 10.12; 0, 25.11.

The aqueous mother liquors were concentrated in vacuo to give a solidwhich was recrystallized from acetone; it consisted of methylN-acetyl-3,4-O-isopropylidene-ot-thiolincosaminide of melting point178180 C. and rotation [u] +l89 (c., 0.5137, water).

B. CONVERSION OF THE OXAZOLINE DERIVED FROM METHYLN-ACETYL-3,4-O-ISOPROPYLIDENE a-THIO- LINCOSAMINIDE TO METHYLN-ACETYL-3,4-O ISO- PROPYLIDE NE-wTHI O-LINCOSAMLNIDE A solution wasprepared of the oxazoline derived from methylN-acetyl-3,4-O-isopropylidene-a-thiolincosaminide in ml. of hot waterand heated under reflux for 2 hours at which time thin-layerchromatography on silica gel revealed the conversion of the startingmaterial to methyl N-acetyl-3,4-O-isopropylidene-a-thiolincosaminide.The water was then removed in vacuo at 40 C., leaving a colorlesscrystalline solid which was recrystallized from a small volume of hotwater to give colorless needles of methylN-acetyl-3,4-O-isopropylidene-a-thiolincosaminide of melting point178180 C. and rotation [a] +190 (c., 1.223, water).

0. METHYLATION OF METHYL N-ACETYL-3,4O-ISO-PROPYLIDENE-a-THIOLLNCOSAMI'NIDE Potassium metal (1.16 g.) was dissolvedin 100 ml. of t-butyl alcohol (previously dried over sodium metal) understirring and reflux. The solvent was removed as completely as possibleby distillation and finally by distillation in a vacuum of 15 mm. Hg. Tothe dry, solid residue was added 100 ml. of dry benzene, which wasremoved by distillation to leave a fine powder. This was treated againwith benzene which was distilled off to insure complete removal oftert-butyl alcohol.

To the thus-obtained, powdery potassium tert-butoxide was added 200 ml.of dry benzene and the mixture was stirred magnetically at roomtemperature until an opalescent suspension resulted. To this was added 5g. of methyl N-acetyl-3,4-0-isopropylidene-a-thiolincosaminide and themixture was then stirred overnight at room temperature at the end ofwhich time all of the solid had dissolved.

To this mixture was added 42.4 iodide and the mixture was stirred atroom temperature for 1 /2 hours; after 1 hour, the mixture gave aneutral reaction with moist pH paper. The reaction mixture was filteredto remove potassium iodide and the potassium iodide was washed withbenzene; the benzene was added to the filtrate. The filtrate andwashings were distilled in vacuo at C. to give a colorless syrup whichwas subjected to counter current distribution in the system ethylacetatezethanolzwater in the ratio 4: 1:2. After 500 transfers the threecomponents, as indicated by thin-layer chromatography, had beencompletely resolved. The major g. (18.6 ml.) of methyl components weremethyl N-acetyl-3,4-O -isopropylidene- 7-O-methyl-u-thiolincosaminideand methyl N-acetyl-3,4- O isopropylidene-Z-O-methyl athiolincosaminide. A minor component was methylN-acetyl-Hi,4-O-isopropylidene-Z,7-di-O-methyl-a-thiolincosaminide.

Removal of solvents from combinedtubes No. 250 310 (K=1.30) inclusivegave a glassy material which crystallized from ethyl acetate:SkellysolveB hexanes to yield methylN-acetyl-3,4-O-isopropylidene-2-O-methyla-thiolincosaminide as short,colorless prisms of melting point 176-177" C. and rotation [a] +l76 (c.,0.6220, chloroform) Analysis.Calcd. for C H NO S: C, 51.57; H, 7.79; N,4.01; S, 9.17; OMe, 8.88. Found: C, 51.82; H, 8.10; N, 4.08; S, 8.94;OMe, 8.49.

Removal of solvents from combined tubes No. 330- 384 (K=2.52) inclusiveby distillation gave a glassy product which crystallized slowly onstanding. Recrystallization from ether gave clusters of minute,colorless needles of methylN-acetyl-7-O-methyl-3,4-O-isopropylidene-ot-thiolincosaminide.

Similarly, evaporation of the solvents from combined tubes No. 410450(K=5.67) gave a colorless glassy product. Recrystallization from ethergave colorless, chunky needles of methyl N-acetyl-Z,7-di-O-methyl-3,4-O-isopropylidene-u-thiolincosaminide of melting point 124.5126 C. androtation [a] +l84 (c., 0.8390, chloroform) Analysis.-Calcd. for C H NOS: C, 52.88; H, 8.04; N, 3.85; S, 8.82; OCH 17.08. Found: C, 53.02; H,7.95; N, 4.05; S, 8.73; OCH 15.92.

c. METHYL N-ACETYL-2-O-METHYL-a-THIOLIN- CO SAMINIDE A mixture of 2 g.of methyl N-acetyl-2-O-methyl-3,4-

' O-isopropylidene-u-thiolincosaminide in 50 ml. of 0.25

N hydrochloric acid was magnetically stirred at room temperature (about25 C.). The solid starting material dissolved within minutes. After 1%hours, thin-layer chromatography showed the complete absence of startingmaterial.

The strongly acidic solution was stirred with a polystyrene quaternaryammonium anion exchange resin until the colorless supernatant solutiongave a neutral reaction with pH paper. Filtration, washing of the resinwith water, and removal of the water from the combined filtrate andwashes in vacuo gave 1.68 g. of a colorless crystalline residue whichwas crystallized from methanol-ether to give long, colorless, feltedneedles of methyl N-acetyl-2-O-methyl-a-thiolincosaminide of meltingpoint 237-238 C.

Analysis.Calcd. for C H NO S: I, 46.56; H, 7.49; N, 4.53; S, 10.36.Found: C, 46.72; H, 7.44; N, 4.37; S, 10.34.

D. METHYL '2-O-METHYL-a-THIOLINCOSAMINIDE The thus-obtained methylN-acetyl-Z-O-methyl-a-thiolincosaminide was heated with 6 ml. ofhydrazine hydrate under reflux for a period of 22 hours. The excesshydrazine hydrate was removed by distillation in vacuo and the residuewas three times recrystallized from ethanol-water to give methyl2-O-methyl-a-thiolincosaminide.

E. METHYL 7'O-METHYL-a-THIOLINCOSAMINIDE In the manner given in Parts Cand D methyl N- acetyl 7 0methyl-3,4-O-isopropylidine-a-thiolincosaminide was hydrolyzed and thenhydrazinolyzed to give methyl 7-O-methyl-a-thiolincosaminide.

F. METHYL 2,7-DI-O-METHYL-a-THIOLINCOSAMI-NIDE In the manner given inParts C and D, methyl N- acetyl 2,7 diO-methyl-3,4-O-isopropylidene-a-thiolincosaminide was hydrolyzed andthen hydrazinolyzed to give methyl 2,7-di-O-methyl-a-thiolincosaminide.

By substituting the methyl a-thiolincosaminide by methylfi-thiolincosaminide or by the other alkyl 11- or fi-thiolincosaminidesgiven above, the corresponding alkyl sees 3]. 2 0,-, 70-, and2,7-di-O-alkyl O6- and fi-thiolincosaminides are obtained. v

Following the procedure of Example 1, compounds of the formula:

wherein X R or Z as given above, R is as given above, and one of R and Ris alkyl of not more than 12 carbon atoms and the other is hydrogen oralkyl of not more than 12 carbon atoms, are obtained.

By substituting the methyl N-acetyl-or-thiolincosaminide by2-hydroxyethyl ocand B-thiolincosaminides, the corresponding alkoxyethylN-acetyl-2-O-, 7-0-, and 2,7-di- O-alkyl-aand fl-thiolincosaminides areobtained. By substituting the methyl N-acetyl-a-thiolincosaminide by 2-trityloxyethyl N16tylocand ti-thiolincosaminides, the corresponding2-trityloxyethy1 N-acetyl-2-O-, 7-0-, and 2,7-di-O-alkyl-or-andfi-thiolincosaminides are obtained. By removing the trityl group byhydrolyzing with 80% aqueous acetic acid and the N-acetyl byhydrazinolysis, the corresponding 2-hydroxyethyl 2-0-, 7-0-, and 2,7-di-O-aIkyl-aand fl-thiol'incosaminides are obtained.

Following the procedures of Example 1, compounds of the formula:

wherein X:R or Z as given above, R and R are as given above, and atleast one of R and R is alkyl of not more than 12 carbon atoms and theother is hydrogen or alkyl of not more than 12 carbon atoms, areobtained.

Example 11.-Meth.yl 6-amino-6,8-dide0xy-L-thre0-a-D-galacto-octopyrcmoside HzN- Ho i\ OH Sale OH LI A.3,4-0-1SQPROPYLIDENELINCOMYCIN CH N 3 Cal-l H l N LII o O SCHa Asolution of 9.8 g. of lincomycin in 150 ml. of acetone is added to asolution of 9.8 g. of p-toluenesulfonic acid monohydrate in 100 ml. ofacetone with good stirring and avoidance of exposure to moisture. Themixture is stirred at ambient temperature for 1 hour, after which 100ml.

XLIX

of anhydrous ether is-added and stirring is continued in an ice-bath for0.5 hour, The mixture is filtered and the solid is dried in vacuo at 50C.; yield 13.35 g. (85.5%) of 3,4-O-isopropylidenelincomycinp-toluenesulfonate. An additional 1.15 g. (7.4%) can be recovered fromthe mother liquors by adding 350 ml. of anhydrous ether to the motherliquor from the previous filtering operation and chilling the solutionfor 1 hour. The 14.5 g. so obtained are suspended in 260 ml. of etherand shaken vigorously with ml. of 5% potassium bicarbonate solution. Theaqueous layer is back-extracted with two lGO-ml. portions of ether. Theether, extracts are Washed with 50 ml. of saturated sodium chloridesolution and then filtered through anhydrous sodium sulfatewThe ether isevaporated under vacuum, leaving 7.9 g. (73.1%) of3,4-O-isopropylidenelincomycin which is dissolved in 25 ml. of ethylacetate and concentrated to about 10 tolSml. The concentrate is allowedto stand at room temperature for several hours and then refrigeratedovernight. The crystals are filtered from the solution and washedsparingly with cold ethyl acetate; yield 4.55 g. (42.2%) of3,4-O-isopropylidenelincomycin having a rneltingpoint of 126 128 C., andan optical rotation of [m] |lOllO2 (c., 1, methylene chloride). B. 7DEHYDRO-3A-IS-OPROPYLIDENE LI'NCOMYCIN To a solution of 6 g. (0.0135mole) of isopropylidenelincomyein in 75 ml. of pyridine added 12 g.(excess) chromic oxide. The solution warms up about 20 C. After one hourthe mixture was added to a solution containing 250 ml. each of ethylether and ethyl acetate. This was then filtered and evaporated to asyrup, 8.4 g. This syrup was distributed in a SOD-transfer countercurrent distribution using the system, waterzethylacetatezethanolzcyclohexane (1: l: 1: 1).7-dehydro-3,4-isopropylidenelincomycin was isolated as the peak fractionfrom tubes 330 380, 14:2.45.

- Analysis.-Calcd. for C H N O SI C. 56.72; H, 8.16; N, 6.30; S, 7.21.Found: C, 56.37; H, 7.62; N, 6.51; S, 6.84.

C. 3,4-ISOPROPYLIDENEEPILINCOMYCIN To 1.6 g. of Craigpure7-dehydro-isoprpylidenelincomycin in 75 ml. of methanol, was added 400mg. of sodium borohydride. After 1.5 hr. this solution was evaporated todryness on a rotary evaporator. The residue was added to 25 ml. of waterand extracted three times with 25 ml. each of methylene chloride. Theextract was back-washed with 15 ml. of Water, then dried over magnesiumchloride. The residue, 1.4 g., was distributed in a SOD-transfer countercurrent distribution using the solvent system, water:ethylacetatezethanohcyclohexane (1:1:1:1) and a single peak which fit thetheoretical was observed at K:1.05. The material in tubes 240 to 28 0'was isolated as a syrup. I

Analysis. Calcd. for C H N O S: C, 56.47; H, 8.58; N, 6.27; S, 7.13.Found: C, 56.24; H, 8.54; N, 6.13; S, 7.01.

Thin layer chromatography (TLC) showed that this material consisted oftwo substances, one of which was 3,4-isopropylidenelincomycin.. Theother, 3,4-isopropylideneepilincornycin, moved slightly slower.

D. EPILINCOMYCIN room temperof 0.25 N hydrochloric acid and 40 ml. ofethanol. It was then kept at C. for 4 days. Following neutralizationwith sodium bicarbonate, it was evaporated to 25 ml., then extractedwith chloroform. The extract was washed with a little water and driedover magnesium sulfate, then evaporated to a residue. Thin layerchromatography of the residue showed two substances, both of which wereactive against S. lutea. The residue was chromatographed on a 14 X A"Florisil (a synthetic silicate of the type described in U.S. Patent2,393,625) column which was eluted gradient ly with solvent which variedcontinuously from 100% Skellysolve B (technical hexane) to 100% acetone.The total volume was 5000 ml. The two compounds were thus separated.

Fraction I: Tubes 53-65 Assay 450 meg/ml.

Analysis.-Calcd. for C18H34N206S1 C, H, N, 6.60; S, 7.56. Found: C,50.19; H, 7.91; N, 6.05; S, 6.42.

Fraction II: Tubes 73-104. Lincomycin. Assay 950 meg/mg.

(40 ml. cuts). Epilincomycin.

E. METHYL EPI-a-THIO'LI'NCOS-AMINIDE Following the hydrazinolyticprocedure of Example 1, Part B, epilincomycin is converted to methylepi-ot-thiolincosaminide.

In place of lincomycin there can be substituted methyl-N-acetyl-a-thiolincosaminide. The process can also be applied to other6-acylamino-6,8-dideoxy-D-erythro-D-galacto-octopyranose compounds ofthe formula:

0 R L IV where Ac, Y and R are as given above to form the correspondingepimers:

which can be alkylated by the procedure given above to provide compoundsof the formula:

0R LVI There can thus be obtained 6-amino-6,S-dideoxy-L-threo-D-galacto-; 6-amino-2-O-alkyl-6,8-dideoxy L threo D- galacto-;6-amino-7-O-alkyl-6,8-dideoxy-L-threo-D-galacto-; and6-amino-2,7-di-O-alkyl-6,8-dideoxy-L threo D- galacto-octopyranosecompounds which in turn can be treated by the procedure of Example 1,Part C et seq., to give compounds of the formula:

LVII

wherein X=Z or R as are as given above.

The intermediates of Formula III can be used as a buffer or as anantacid. They react with isocyanates to form urethanes and can be usedto modify polyurethane resins. The thiocyanic acid addition salt whencondensed with formaldehyde forms resinous materials useful as picklinginhibitors according to U.S. Patents 2,425,320 and 2,606,- 155, The freebases also make good vehicles for toxic acids. For example, thefluosilicic acid addition salts are useful as mothproofing agentsaccording to U.S. Patents 1,915,- 334 and 2,075,359 and thehexafluoroarsenic acid and hexafluorophosphoric acid addition salts areuseful as parasiticides according to U.S. Patents 3,122,536 and3,122,552.

Various acid-addition salts of the free base form of the intermediatesfor Formula III can be made by neutralizing the free base with theappropriate acid to below about pH 7.0, and advantageously to about pH 2to pH 6. Suitable acids for this purpose include hydrochloric, sulfuric,phosphoric, thiocyanic, fluosilicic, hexafluoroarsenic,hexafluorophosphoric, acetic, succinic, citric, lactic, maleic, fumaric,pamoic, cholic, palmitic, mucic, camphoric, glutaric, glycolic,phthalic, tartaric, lauric, stearic, salicyclic, 3-phenylsalicylic,S-phenylsalicylic, 3-methylglutaric, orthosulfobenzoic,cyclohexanesulfamic, cyclopentanepropionic, l,2-cyclohexanedicarboxylic,4-cyclohexenecarboxylic, octadecenylsuccinic, octenylsuccinic,methanesulfonic, benzenesulfonic, helianthic, Reineckes,dimethyldithiocarbamic, cyclohexylsulfamic, hexadecylsulfamic,octadecylsulfamic, sorbic, monochloroacetic, undecylenic,4'-hydroxyazobenzene-4-sulfonic, octayldecylsulfuric, picric, benzoic,cinnamic, and like acids.

The acid addition salts can be used for the same purposes as the freebase or they can be employed to upgrade the same. For example, the freebase can be converted to an insoluble salt, such as the picrate, whichcan be subjected to purification procedures, for example, solventextractions and washings, chromatography, fractional liquidliquidextractions, and crystallization and then used to regenerate the freebase form by treatment with alkali or to make a different salt bymetathesis. Or the free base can be converted to a water-soluble salt,such as the hydrochloride or sulfate and the aqueous solution of thesalt extracted with various water-immiscible solvents beforeregenerating the free base form by treatment of the thusgiven above andY, R, R and R extracted acid solution or converted to another salt by 1Q XXXVII, XXXVIII, XLV, and XLVII.

TABLE I CH3 H CH3 C 115 H CH3 H CH3 CH2 H C 115 H CH3 CH3 H CH3 C113 H C115 H H CH; 11

Typical compounds prepared according to Examples 5, 6, and 7 includeboth the cis and trans forms of the following compounds according toFormulas XL, XLII,

Typical compounds according to Formulas XLIX and L prepared according toExample 10 include each of the compounds listed above in which theZ-O-hydrogen (R and the 7-O-hydrogen (R and the 2-0- and 7-O-hydrogens(R and R are replaced by alkyl, for example, methyl, ethyl, propyl,butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, undecyl, and dodecyland the isomeric forms thereof.

Typical compounds prepared according to Formula LVII include each of theabove compounds in the L- threo configuration.

We claim:

1. A compound of the formula or the formula:

OHa

Rio-- wherein Z is a protective group removable by hydrogenolysis; R isalkylidene of not more than 20 carbon atoms, cycloalkylidene of 3 to notmore than 8 carbon atoms, or aralkylidene of not more than 12 carbonatoms; R and R are hydrogen or alkyl of not more than 12 carbon atoms;and R is hydrogen, alkyl of not more than 12 carbon atoms, salicyloyl,fatty acid acyl of not more than 36 18 carbon atoms, and acrylic acidacyl of not more than 18 carbon atoms.

2. A compound of the formula:

Z CH a l/ lk 31 o ll/ OR: (N)

wherein R, R R and Z are as given in claim 1.

3. A compound of the formula Z CH3 N K \I C-NH 20 r" l OH SCH on -o R \ll/ 2 2 r v 033 (Q) or the formula:

2 liq CH3 Rio-- l -0 RH O HO \\SCH2CHz-OR4 3,

1 (LR; wherein R, R R R and Z are as given in claim 1.

4. A compound of the formula: Z 1|I CH;

Rio-- T iTNH" --0 5 11 RH O HO i \OH l r i 0R: (S) wherein R, R R and Zare as given in claim 1.

5. A member of the group consisting of the free base and acid-additionsalt forms of a compound of the formula:

' (H3 N l mo l C-NH H EH 0 HO( \l l/ SGHzOHz-O-Jh or the formula:

I a N 1/ 1 ONH L H0 somom-o-rn l iH 1 CR3 (W) 7 wherein R, R R and R areas given in claim 1.

6. A member of the group consisting of the free base and acid-additionsalt forms of a compound of the formula:

l RzO- 34,,

wherein R, R and R are as given in claim 1.

7. A member of the group consisting of the free base and acid-additionsalt forms of a compound of the forwherein R, R R and R are as given inclaim 1 and R is alkyl of not more than 20 carbon atoms, cycloalkyl ofnot more than 8 carbon atoms, and aralkyl of not more than 12 carbonatoms.

8. A member of the group consisting of the free base and acid-additionsalt forms of a compound of the formula:

i 11 CH3 R20 I (LR, (AB) wherein R, R and R are as given in claim 14 andR is alkyl of not more than 20 carbon atoms, cycloalkyl of not more than8 carbon atoms, and aralkyl of not more than 12 carbon atoms.

9. A member of the group consisting of the free base and acid-additionsalt forms of a compound of the formula:

I one N IK I fiNH-- E II II RH O OH somom-o-lu wherein alkyl is of notmore than 12 carbon atoms, R is a member of the group consisting ofalkylidene of not more than 20 carbon atoms, cycloalkylidene of 3 to notmore than 8 carbon atoms, and aralkylidene of not more than 12 carbonatoms, R is a member of the group consisting of hydrogen, alkyl of notmore than 20 carbon atoms, cycloalkyl of 3 to not more than 8 carbonatoms, and aralkyl of not more than 12 carbon atoms, R; is hydrogen,alkyl of not more than 12 carbon atoms, salicyloyl, fatty acid acyl ofnot more than 18 carbon atoms, and acrylic acid acyl of not more than 18carbon atoms; and one of R and R is an alkyl group of not more than 12carbon atoms and the other is a member of the group consisting ofhydrogen and an alkyl of not more than 12 carbon atoms.

10. A process which comprises desulfurizing with mercuric chloride acompound of the formula:

and acylating the resulting product to form a compound of the formula:

brominating with HBr to form a compound of the formula:

alkylating with a compound of the formula R -halogen to form a compoundof the formula:

and R wherein R is alkyl of not more than 12 carbon atoms.

11. A member of the group consisting of the free base wherein R is asgiven in claim 10.

12. A process which comprises acylating of the formula:

a compound wherein alkyl is methyl or ethyl with thiobenzoyl halide toform the tetrathionobenzoate, brominating the tetrathionobenzoate toform a compound of the formula:

heating the resulting compound in acetone in the absence of water toform a compound of the formula:

alkylating the resulting compound with a compound of the formula:

R -halogen in the presence of mild base to form a compound of theformula:

Oso-

AcNH- oso K 6666 1 w O 0 O Ac being a member of the group consisting ofalkanoyl and aralkanoyl of not more than 12 carbon atoms, n is aninteger of 1 to 2, and R is a member of the group consisting of --CH CHOH, -CH CH OR wherein R is alkyl of not more than 12 carbon atoms, andalkyl of not more than 12 carbon atoms other than the alkyl of formula(BC). 7

13. The process of claim 12 wherein the compound of formula BF issubjected to hydrazinolysis to form a compound of the formula:

HzN- k 31 011 \l 310 wherein R is as given in claim 12.

14. A member of the group consisting of the free base and acid-additionsalt forms of a compound of the for- 0R (BI) wherein R and R are membersof the group consisting of hydrogen and alkyl of not more than 12 carbonatoms.

References Cited UNITED STATES PATENTS 3,255,180 6/1966 Hoeksema26O--210 LEWIS GOTTS, Primary Examiner.

I. R. BROWN, Assistant Examiner.

1. A COMPOUND OF THE FORMULA